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Sample records for streptococcus pyogenes causantes

  1. Streptococcus pyogenes in Human Plasma

    PubMed Central

    Malmström, Johan; Karlsson, Christofer; Nordenfelt, Pontus; Ossola, Reto; Weisser, Hendrik; Quandt, Andreas; Hansson, Karin; Aebersold, Ruedi; Malmström, Lars; Björck, Lars

    2012-01-01

    Streptococcus pyogenes is a major bacterial pathogen and a potent inducer of inflammation causing plasma leakage at the site of infection. A combination of label-free quantitative mass spectrometry-based proteomics strategies were used to measure how the intracellular proteome homeostasis of S. pyogenes is influenced by the presence of human plasma, identifying and quantifying 842 proteins. In plasma the bacterium modifies its production of 213 proteins, and the most pronounced change was the complete down-regulation of proteins required for fatty acid biosynthesis. Fatty acids are transported by albumin (HSA) in plasma. S. pyogenes expresses HSA-binding surface proteins, and HSA carrying fatty acids reduced the amount of fatty acid biosynthesis proteins to the same extent as plasma. The results clarify the function of HSA-binding proteins in S. pyogenes and underline the power of the quantitative mass spectrometry strategy used here to investigate bacterial adaptation to a given environment. PMID:22117078

  2. [Streptococcus pyogenes pathogenic factors].

    PubMed

    Bidet, Ph; Bonacorsi, S

    2014-11-01

    The pathogenicity of ß-hemolytic group A streptococcus (GAS) is particularly diverse, ranging from mild infections, such as pharyngitis or impetigo, to potentially debilitating poststreptococcal diseases, and up to severe invasive infections such as necrotizing fasciitis or the dreaded streptococcal toxic shock syndrome. This variety of clinical expressions, often radically different in individuals infected with the same strain, results from a complex interaction between the bacterial virulence factors, the mode of infection and the immune system of the host. Advances in comparative genomics have led to a better understanding of how, following this confrontation, GAS adapts to the immune system's pressure, either peacefully by reducing the expression of certain virulence factors to achieve an asymptomatic carriage, or on the contrary, by overexpressing them disproportionately, resulting in the most severe forms of invasive infection.

  3. Streptococcus pyogenes adhesion and colonization.

    PubMed

    Brouwer, Stephan; Barnett, Timothy C; Rivera-Hernandez, Tania; Rohde, Manfred; Walker, Mark J

    2016-11-01

    Streptococcus pyogenes (group A Streptococcus, GAS) is a human-adapted pathogen responsible for a wide spectrum of disease. GAS can cause relatively mild illnesses, such as strep throat or impetigo, and less frequent but severe life-threatening diseases such as necrotizing fasciitis and streptococcal toxic shock syndrome. GAS is an important public health problem causing significant morbidity and mortality worldwide. The main route of GAS transmission between humans is through close or direct physical contact, and particularly via respiratory droplets. The upper respiratory tract and skin are major reservoirs for GAS infections. The ability of GAS to establish an infection in the new host at these anatomical sites primarily results from two distinct physiological processes, namely bacterial adhesion and colonization. These fundamental aspects of pathogenesis rely upon a variety of GAS virulence factors, which are usually under strict transcriptional regulation. Considerable progress has been made in better understanding these initial infection steps. This review summarizes our current knowledge of the molecular mechanisms of GAS adhesion and colonization. © 2016 Federation of European Biochemical Societies.

  4. Genetic Manipulation of Streptococcus pyogenes (The Group A Streptococcus, GAS)

    PubMed Central

    Le Breton, Yoann; McIver, Kevin S.

    2013-01-01

    Streptococcus pyogenes (the group A streptococcus, GAS) is a Gram-positive bacterium responsible for a wide spectrum of diseases ranging from mild superficial infections (pharyngitis, impetigo) to severe often life-threatening invasive diseases (necrotizing fasciitis, streptococcal toxic shock syndrome) in humans. This unit describes molecular techniques for the genetic manipulation of S. pyogenes with detailed protocols for transformation, gene disruption, allelic exchange, transposon mutagenesis, and genetic complementation. PMID:24510894

  5. Genetic manipulation of Streptococcus pyogenes (the Group A Streptococcus, GAS).

    PubMed

    Le Breton, Yoann; McIver, Kevin S

    2013-10-02

    Streptococcus pyogenes (the Group A Streptococcus, GAS) is a Gram-positive bacterium responsible for a wide spectrum of diseases ranging from mild superficial infections (pharyngitis, impetigo) to severe, often life-threatening invasive diseases (necrotizing fasciitis, streptococcal toxic shock syndrome) in humans. This unit describes molecular techniques for the genetic manipulation of S. pyogenes with detailed protocols for transformation, gene disruption, allelic exchange, transposon mutagenesis, and genetic complementation.

  6. Murine models of Streptococcus pyogenes infection.

    PubMed

    Roberts, Samantha; Scott, June R; Husmann, Linda K; Zurawski, Christine A

    2006-09-01

    This unit describes procedures for testing virulence of Streptococcus pyogenes in mice. S. pyogenes is an important human pathogen and causes one of the most common childhood diseases. The syndromes that result from S. pyogenes infection are diverse, ranging from mild, superficial throat or skin infection to severe, invasive disea/se that is often lethal. Thus, a greater understanding of the virulence factors of this bacterium and development of modalities to prevent or relieve the infections it causes are important. Since S. pyogenes is a strictly human pathogen (with the exception of a single strain), the value of all animal models is limited. This unit describes a model for long-term throat colonization following the natural route of infection (inhalation), one for pneumonia and systemic dissemination following intratracheal inoculation, and one for systemic dissemination following the more natural route of skin infection. In addition, methods are presented for culturing S. pyogenes from tissues of the infected animal.

  7. Antimicrobial Drug Use and Macrolide-Resistant Streptococcus pyogenes, Belgium

    PubMed Central

    Van Heirstraeten, Liesbet; Coenen, Samuel; Lammens, Christine; Hens, Niel; Goossens, Herman

    2012-01-01

    In Belgium, decreasing macrolide, lincosamide, streptogramins B, and tetracycline use during 1997–2007 correlated significantly with decreasing macrolide-resistant Streptococcus pyogenes during 1999–2009. Maintaining drug use below a critical threshold corresponded with low-level macrolide-resistant S. pyogenes and an increased number of erm(A)-harboring emm77 S. pyogenes with low fitness costs. PMID:22932671

  8. Gene Repertoire Evolution of Streptococcus pyogenes Inferred from Phylogenomic Analysis with Streptococcus canis and Streptococcus dysgalactiae

    PubMed Central

    Lefébure, Tristan; Richards, Vince P.; Lang, Ping; Pavinski-Bitar, Paulina; Stanhope, Michael J.

    2012-01-01

    Streptococcus pyogenes, is an important human pathogen classified within the pyogenic group of streptococci, exclusively adapted to the human host. Our goal was to employ a comparative evolutionary approach to better understand the genomic events concomitant with S. pyogenes human adaptation. As part of ascertaining these events, we sequenced the genome of one of the potential sister species, the agricultural pathogen S. canis, and combined it in a comparative genomics reconciliation analysis with two other closely related species, Streptococcus dysgalactiae and Streptococcus equi, to determine the genes that were gained and lost during S. pyogenes evolution. Genome wide phylogenetic analyses involving 15 Streptococcus species provided convincing support for a clade of S. equi, S. pyogenes, S. dysgalactiae, and S. canis and suggested that the most likely S. pyogenes sister species was S. dysgalactiae. The reconciliation analysis identified 113 genes that were gained on the lineage leading to S. pyogenes. Almost half (46%) of these gained genes were phage associated and 14 showed significant matches to experimentally verified bacteria virulence factors. Subsequent to the origin of S. pyogenes, over half of the phage associated genes were involved in 90 different LGT events, mostly involving different strains of S. pyogenes, but with a high proportion involving the horse specific pathogen S. equi subsp. equi, with the directionality almost exclusively (86%) in the S. pyogenes to S. equi direction. Streptococcus agalactiae appears to have played an important role in the evolution of S. pyogenes with a high proportion of LGTs originating from this species. Overall the analysis suggests that S. pyogenes adaptation to the human host was achieved in part by (i) the integration of new virulence factors (e.g. speB, and the sal locus) and (ii) the construction of new regulation networks (e.g. rgg, and to some extent speB). PMID:22666370

  9. Thermoregulation of Capsule Production by Streptococcus pyogenes

    PubMed Central

    Kang, Song Ok; Wright, Jordan O.; Tesorero, Rafael A.; Lee, Hyunwoo; Beall, Bernard; Cho, Kyu Hong

    2012-01-01

    The capsule of Streptococcus pyogenes serves as an adhesin as well as an anti-phagocytic factor by binding to CD44 on keratinocytes of the pharyngeal mucosa and the skin, the main entry sites of the pathogen. We discovered that S. pyogenes HSC5 and MGAS315 strains are further thermoregulated for capsule production at a post-transcriptional level in addition to the transcriptional regulation by the CovRS two-component regulatory system. When the transcription of the hasABC capsular biosynthetic locus was de-repressed through mutation of the covRS system, the two strains, which have been used for pathogenesis studies in the laboratory, exhibited markedly increased capsule production at sub-body temperature. Employing transposon mutagenesis, we found that CvfA, a previously identified membrane-associated endoribonuclease, is required for the thermoregulation of capsule synthesis. The mutation of the cvfA gene conferred increased capsule production regardless of temperature. However, the amount of the capsule transcript was not changed by the mutation, indicating that a post-transcriptional regulator mediates between CvfA and thermoregulated capsule production. When we tested naturally occurring invasive mucoid strains, a high percentage (11/53, 21%) of the strains exhibited thermoregulated capsule production. As expected, the mucoid phenotype of these strains at sub-body temperature was due to mutations within the chromosomal covRS genes. Capsule thermoregulation that exhibits high capsule production at lower temperatures that occur on the skin or mucosal surface potentially confers better capability of adhesion and invasion when S. pyogenes penetrates the epithelial surface. PMID:22615992

  10. Thermoregulation of capsule production by Streptococcus pyogenes.

    PubMed

    Kang, Song Ok; Wright, Jordan O; Tesorero, Rafael A; Lee, Hyunwoo; Beall, Bernard; Cho, Kyu Hong

    2012-01-01

    The capsule of Streptococcus pyogenes serves as an adhesin as well as an anti-phagocytic factor by binding to CD44 on keratinocytes of the pharyngeal mucosa and the skin, the main entry sites of the pathogen. We discovered that S. pyogenes HSC5 and MGAS315 strains are further thermoregulated for capsule production at a post-transcriptional level in addition to the transcriptional regulation by the CovRS two-component regulatory system. When the transcription of the hasABC capsular biosynthetic locus was de-repressed through mutation of the covRS system, the two strains, which have been used for pathogenesis studies in the laboratory, exhibited markedly increased capsule production at sub-body temperature. Employing transposon mutagenesis, we found that CvfA, a previously identified membrane-associated endoribonuclease, is required for the thermoregulation of capsule synthesis. The mutation of the cvfA gene conferred increased capsule production regardless of temperature. However, the amount of the capsule transcript was not changed by the mutation, indicating that a post-transcriptional regulator mediates between CvfA and thermoregulated capsule production. When we tested naturally occurring invasive mucoid strains, a high percentage (11/53, 21%) of the strains exhibited thermoregulated capsule production. As expected, the mucoid phenotype of these strains at sub-body temperature was due to mutations within the chromosomal covRS genes. Capsule thermoregulation that exhibits high capsule production at lower temperatures that occur on the skin or mucosal surface potentially confers better capability of adhesion and invasion when S. pyogenes penetrates the epithelial surface.

  11. The Streptococcus pyogenes proteome: maps, virulence factors and vaccine candidates

    PubMed Central

    Dmitriev, Alexander V; Chaussee, Michael S

    2011-01-01

    Streptococcus pyogenes is an important cause of human morbidity and mortality worldwide. A wealth of genomic information related to this pathogen has facilitated exploration of the proteome, particularly in response to environmental conditions thought to mimic various aspects of pathogenesis. Proteomic approaches are also used to identify immunoreactive proteins for vaccine development and to identify proteins that may induce autoimmunity. These studies have revealed new mechanisms involved in regulating the S. pyogenes proteome, which has opened up new avenues in the study of S. pyogenes pathogenesis. This article describes the methods used, and progress being made towards characterizing the S. pyogenes proteome, including studies seeking to identify potential vaccine candidates. PMID:21073313

  12. Is Streptococcus pyogenes resistant or susceptible to trimethoprim-sulfamethoxazole?

    PubMed

    Bowen, Asha C; Lilliebridge, Rachael A; Tong, Steven Y C; Baird, Robert W; Ward, Peter; McDonald, Malcolm I; Currie, Bart J; Carapetis, Jonathan R

    2012-12-01

    Streptococcus pyogenes is commonly believed to be resistant to trimethoprim-sulfamethoxazole (SXT), resulting in reservations about using SXT for skin and soft tissue infections (SSTI) where S. pyogenes is involved. S. pyogenes' in vitro susceptibility to SXT depends on the medium's thymidine content. Thymidine allows S. pyogenes to bypass the sulfur-mediated inhibition of folate metabolism and, historically, has resulted in apparently reduced susceptibility of S. pyogenes to sulfur antibacterials. The low thymidine concentration in Mueller-Hinton agar (MHA) is now regulated. We explored S. pyogenes susceptibility to SXT on various media. Using two sets of 100 clinical S. pyogenes isolates, we tested for susceptibility using SXT Etests on MHA containing defibrinated horse blood and 20 mg/liter β-NAD (MHF), MHA with sheep blood (MHS), MHA alone, MHA with horse blood (MHBA), and MHA with lysed horse blood (MHLHBA). European Committee on Antibacterial Susceptibility Testing (EUCAST) breakpoints defined susceptibility (MIC, ≤ 1 mg/liter) and resistance (MIC, >2 mg/liter). In study 1, 99% of S. pyogenes isolates were susceptible to SXT on MHA, MHBA, and MHLHBA, with geometric mean MICs of 0.04, 0.04, and 0.05 mg/liter, respectively. In study 2, all 100 S. pyogenes isolates were susceptible to SXT on MHF, MHS, MHA, and MHLHBA with geometric mean MICs of 0.07, 0.16, 0.07, and 0.09 mg/liter, respectively. This study confirms the in vitro susceptibility of S. pyogenes to SXT, providing support for the use of SXT for SSTIs. A clinical trial using SXT for impetigo is ongoing.

  13. Is Streptococcus pyogenes Resistant or Susceptible to Trimethoprim-Sulfamethoxazole?

    PubMed Central

    Lilliebridge, Rachael A.; Tong, Steven Y. C.; Baird, Robert W.; Ward, Peter; McDonald, Malcolm I.; Currie, Bart J.; Carapetis, Jonathan R.

    2012-01-01

    Streptococcus pyogenes is commonly believed to be resistant to trimethoprim-sulfamethoxazole (SXT), resulting in reservations about using SXT for skin and soft tissue infections (SSTI) where S. pyogenes is involved. S. pyogenes' in vitro susceptibility to SXT depends on the medium's thymidine content. Thymidine allows S. pyogenes to bypass the sulfur-mediated inhibition of folate metabolism and, historically, has resulted in apparently reduced susceptibility of S. pyogenes to sulfur antibacterials. The low thymidine concentration in Mueller-Hinton agar (MHA) is now regulated. We explored S. pyogenes susceptibility to SXT on various media. Using two sets of 100 clinical S. pyogenes isolates, we tested for susceptibility using SXT Etests on MHA containing defibrinated horse blood and 20 mg/liter β-NAD (MHF), MHA with sheep blood (MHS), MHA alone, MHA with horse blood (MHBA), and MHA with lysed horse blood (MHLHBA). European Committee on Antibacterial Susceptibility Testing (EUCAST) breakpoints defined susceptibility (MIC, ≤1 mg/liter) and resistance (MIC, >2 mg/liter). In study 1, 99% of S. pyogenes isolates were susceptible to SXT on MHA, MHBA, and MHLHBA, with geometric mean MICs of 0.04, 0.04, and 0.05 mg/liter, respectively. In study 2, all 100 S. pyogenes isolates were susceptible to SXT on MHF, MHS, MHA, and MHLHBA with geometric mean MICs of 0.07, 0.16, 0.07, and 0.09 mg/liter, respectively. This study confirms the in vitro susceptibility of S. pyogenes to SXT, providing support for the use of SXT for SSTIs. A clinical trial using SXT for impetigo is ongoing. PMID:23052313

  14. CRISPR inhibition of prophage acquisition in Streptococcus pyogenes.

    PubMed

    Nozawa, Takashi; Furukawa, Nayuta; Aikawa, Chihiro; Watanabe, Takayasu; Haobam, Bijaya; Kurokawa, Ken; Maruyama, Fumito; Nakagawa, Ichiro

    2011-05-06

    Streptococcus pyogenes, one of the major human pathogens, is a unique species since it has acquired diverse strain-specific virulence properties mainly through the acquisition of streptococcal prophages. In addition, S. pyogenes possesses clustered regularly interspaced short palindromic repeats (CRISPR)/Cas systems that can restrict horizontal gene transfer (HGT) including phage insertion. Therefore, it was of interest to examine the relationship between CRISPR and acquisition of prophages in S. pyogenes. Although two distinct CRISPR loci were found in S. pyogenes, some strains lacked CRISPR and these strains possess significantly more prophages than CRISPR harboring strains. We also found that the number of spacers of S. pyogenes CRISPR was less than for other streptococci. The demonstrated spacer contents, however, suggested that the CRISPR appear to limit phage insertions. In addition, we found a significant inverse correlation between the number of spacers and prophages in S. pyogenes. It was therefore suggested that S. pyogenes CRISPR have permitted phage insertion by lacking its own spacers. Interestingly, in two closely related S. pyogenes strains (SSI-1 and MGAS315), CRISPR activity appeared to be impaired following the insertion of phage genomes into the repeat sequences. Detailed analysis of this prophage insertion site suggested that MGAS315 is the ancestral strain of SSI-1. As a result of analysis of 35 additional streptococcal genomes, it was suggested that the influences of the CRISPR on the phage insertion vary among species even within the same genus. Our results suggested that limitations in CRISPR content could explain the characteristic acquisition of prophages and might contribute to strain-specific pathogenesis in S. pyogenes.

  15. CRISPR Inhibition of Prophage Acquisition in Streptococcus pyogenes

    PubMed Central

    Aikawa, Chihiro; Watanabe, Takayasu; Haobam, Bijaya; Kurokawa, Ken; Maruyama, Fumito; Nakagawa, Ichiro

    2011-01-01

    Streptococcus pyogenes, one of the major human pathogens, is a unique species since it has acquired diverse strain-specific virulence properties mainly through the acquisition of streptococcal prophages. In addition, S. pyogenes possesses clustered regularly interspaced short palindromic repeats (CRISPR)/Cas systems that can restrict horizontal gene transfer (HGT) including phage insertion. Therefore, it was of interest to examine the relationship between CRISPR and acquisition of prophages in S. pyogenes. Although two distinct CRISPR loci were found in S. pyogenes, some strains lacked CRISPR and these strains possess significantly more prophages than CRISPR harboring strains. We also found that the number of spacers of S. pyogenes CRISPR was less than for other streptococci. The demonstrated spacer contents, however, suggested that the CRISPR appear to limit phage insertions. In addition, we found a significant inverse correlation between the number of spacers and prophages in S. pyogenes. It was therefore suggested that S. pyogenes CRISPR have permitted phage insertion by lacking its own spacers. Interestingly, in two closely related S. pyogenes strains (SSI-1 and MGAS315), CRISPR activity appeared to be impaired following the insertion of phage genomes into the repeat sequences. Detailed analysis of this prophage insertion site suggested that MGAS315 is the ancestral strain of SSI-1. As a result of analysis of 35 additional streptococcal genomes, it was suggested that the influences of the CRISPR on the phage insertion vary among species even within the same genus. Our results suggested that limitations in CRISPR content could explain the characteristic acquisition of prophages and might contribute to strain-specific pathogenesis in S. pyogenes. PMID:21573110

  16. The role of coagulation/fibrinolysis during Streptococcus pyogenes infection

    PubMed Central

    Loof, Torsten G.; Deicke, Christin; Medina, Eva

    2014-01-01

    The hemostatic system comprises platelet aggregation, coagulation and fibrinolysis and is a host defense mechanism that protects the integrity of the vascular system after tissue injury. During bacterial infections, the coagulation system cooperates with the inflammatory system to eliminate the invading pathogens. However, pathogenic bacteria have frequently evolved mechanisms to exploit the hemostatic system components for their own benefit. Streptococcus pyogenes, also known as Group A Streptococcus, provides a remarkable example of the extraordinary capacity of pathogens to exploit the host hemostatic system to support microbial survival and dissemination. The coagulation cascade comprises the contact system (also known as the intrinsic pathway) and the tissue factor pathway (also known as the extrinsic pathway), both leading to fibrin formation. During the early phase of S. pyogenes infection, the activation of the contact system eventually leads to bacterial entrapment within a fibrin clot, where S. pyogenes is immobilized and killed. However, entrapped S. pyogenes can circumvent the antimicrobial effect of the clot by sequestering host plasminogen on the bacterial cell surface that, after conversion into its active proteolytic form, plasmin, degrades the fibrin network and facilitates the liberation of S. pyogenes from the clot. Furthermore, the surface-localized fibrinolytic activity also cleaves a variety of extracellular matrix proteins, thereby enabling S. pyogenes to migrate across barriers and disseminate within the host. This review summarizes the knowledge gained during the last two decades on the role of coagulation/fibrinolysis in host defense against S. pyogenes as well as the strategies developed by this pathogen to evade and exploit these host mechanisms for its own benefit. PMID:25309880

  17. Differentiation of cultured keratinocytes promotes the adherence of Streptococcus pyogenes.

    PubMed Central

    Darmstadt, G L; Fleckman, P; Jonas, M; Chi, E; Rubens, C E

    1998-01-01

    Based on a consideration of the histopathology of nonbullous impetigo that shows localization of Streptococcus pyogenes to highly differentiated, subcorneal keratinocytes, we hypothesized that adherence of an impetigo strain of S. pyogenes would be promoted by terminal differentiation of keratinocytes. An assay was developed in which S. pyogenes adhered via pilus-like projections from the cell wall to the surface of cultured human keratinocytes in a time- and inoculum-dependent manner suggestive of a receptor-mediated process. Terminal differentiation of keratinocytes was induced by increasing the calcium concentration in the growth medium, and was confirmed by morphologic analysis using electron microscopy. Adherence of S. pyogenes was three and fourfold greater to keratinocytes differentiated in 1.0 and 1.5 mM calcium, respectively, compared with undifferentiated keratinocytes in 0.15 mM calcium. The presence of calcium during the adherence assay further enhanced adherence nearly twofold. Adherence occurred preferentially to sites of contact between adjacent keratinocytes, suggesting that the keratinocyte receptor may be a molecule involved in cell-to-cell adhesion. In contrast, nonpathogenic Streptococcus gordonii adhered poorly to keratinocytes regardless of their state of terminal differentiation, and adherence of a pharyngeal strain of S. pyogenes was twofold greater to undifferentiated than differentiated keratinocytes. This is the first report of in vitro adherence of S. pyogenes to keratinocytes in a manner that emulates human impetigo. Adherence of only the impetigo strain, and not the pharyngeal strain of S. pyogenes or the nonpathogenic S. gorgonii isolate, was promoted by keratinocyte differentiation. This result provides a model system for investigating the molecular pathogenesis of streptococcal skin infections. PMID:9421474

  18. In Vitro Antibacterial Activity of Essential Oils against Streptococcus pyogenes.

    PubMed

    Sfeir, Julien; Lefrançois, Corinne; Baudoux, Dominique; Derbré, Séverine; Licznar, Patricia

    2013-01-01

    Streptococcus pyogenes plays an important role in the pathogenesis of tonsillitis. The present study was conducted to evaluate the in vitro antibacterial activities of 18 essential oils chemotypes from aromatic medicinal plants against S. pyogenes. Antibacterial activity of essential oils was investigated using disc diffusion method. Minimum Inhibitory Concentration of essential oils showing an important antibacterial activity was measured using broth dilution method. Out of 18 essential oils tested, 14 showed antibacterial activity against S. pyogenes. Among them Cinnamomum verum, Cymbopogon citratus, Thymus vulgaris CT thymol, Origanum compactum, and Satureja montana essential oils exhibited significant antibacterial activity. The in vitro results reported here suggest that, for patients suffering from bacterial throat infections, if aromatherapy is used, these essential oils, considered as potential antimicrobial agents, should be preferred.

  19. In Vitro Antibacterial Activity of Essential Oils against Streptococcus pyogenes

    PubMed Central

    Sfeir, Julien; Lefrançois, Corinne; Baudoux, Dominique; Derbré, Séverine; Licznar, Patricia

    2013-01-01

    Streptococcus pyogenes plays an important role in the pathogenesis of tonsillitis. The present study was conducted to evaluate the in vitro antibacterial activities of 18 essential oils chemotypes from aromatic medicinal plants against S. pyogenes. Antibacterial activity of essential oils was investigated using disc diffusion method. Minimum Inhibitory Concentration of essential oils showing an important antibacterial activity was measured using broth dilution method. Out of 18 essential oils tested, 14 showed antibacterial activity against S. pyogenes. Among them Cinnamomum verum, Cymbopogon citratus, Thymus vulgaris CT thymol, Origanum compactum, and Satureja montana essential oils exhibited significant antibacterial activity. The in vitro results reported here suggest that, for patients suffering from bacterial throat infections, if aromatherapy is used, these essential oils, considered as potential antimicrobial agents, should be preferred. PMID:23662123

  20. Laboratory Growth and Maintenance of Streptococcus pyogenes (The Group A Streptococcus, GAS)

    PubMed Central

    Gera, Kanika; McIver, Kevin S.

    2013-01-01

    Streptococcus pyogenes is a Gram-positive bacterium that strictly infects humans. It is the causative agent of a broad spectrum of diseases accounting for millions of infections and at least 517, 000 deaths each year worldwide (Carapetis et al., 2005). It is a nutritionally fastidious organism that ferments sugars to produce lactic acid and has strict requirements for growth. To aid in the study of this organism, this unit describes the growth and maintenance of S. pyogenes. PMID:24510893

  1. Fluoride exposure attenuates expression of Streptococcus pyogenes virulence factors.

    PubMed

    Thongboonkerd, Visith; Luengpailin, Jirapon; Cao, Junkai; Pierce, William M; Cai, Jian; Klein, Jon B; Doyle, R J

    2002-05-10

    Fluoridation causes an obvious reduction of dental caries by interference with cariogenic streptococci. However, the effect of fluoride on group A streptococci that causes rheumatic fever and acute poststreptococcal glomerulonephritis is not known. We have used proteomic analysis to create a reference proteome map for Streptococcus pyogenes and to determine fluoride-induced protein changes in the streptococci. Cellular and extracellular proteins were resolved by two-dimensional polyacrylamide gel electrophoresis and identified by matrix-assisted laser desorption ionization mass spectrometry. 183 protein spots were visualized, and 74 spots representing 60 unique proteins were identified. A 16-h exposure to sodium fluoride caused decreased expression of proteins required to respond to cellular stress, including anti-oxidants, glycolytic enzymes, transcriptional and translational regulators, and protein folding. Fluoride caused decreased cellular expression of two well-characterized S. pyogenes virulence factors. Fluoride decreased expression of glyceraldehyde-3-phosphate dehydrogenase, which acts to bind fibronectin and promote bacterial adherence. We also performed proteomic analysis of protein released by S. pyogenes into the culture supernatant and observed decreased expression of M proteins following fluoride exposure. These data provide evidence that fluoride causes decreased expression by S. pyogenes proteins used to respond to stress, virulence factors, and implicated in non-suppurative complications of S. pyogenes, including glomerulonephritis and rheumatic fever.

  2. Factors That Cause Trimethoprim Resistance in Streptococcus pyogenes

    PubMed Central

    Bergmann, René; van der Linden, Mark; Chhatwal, Gursharan S.

    2014-01-01

    The use of trimethoprim in treatment of Streptococcus pyogenes infections has long been discouraged because it has been widely believed that this pathogen is resistant to this antibiotic. To gain more insight into the extent and molecular basis of trimethoprim resistance in S. pyogenes, we tested isolates from India and Germany and sought the factors that conferred the resistance. Resistant isolates were identified in tests for trimethoprim or trimethoprim-sulfamethoxazole (SXT) susceptibility. Resistant isolates were screened for the known horizontally transferable trimethoprim-insensitive dihydrofolate reductase (dfr) genes dfrG, dfrF, dfrA, dfrD, and dfrK. The nucleotide sequence of the intrinsic dfr gene was determined for resistant isolates lacking the horizontally transferable genes. Based on tentative criteria, 69 out of 268 isolates (25.7%) from India were resistant to trimethoprim. Occurring in 42 of the 69 resistant isolates (60.9%), dfrF appeared more frequently than dfrG (23 isolates; 33.3%) in India. The dfrF gene was also present in a collection of SXT-resistant isolates from Germany, in which it was the only detected trimethoprim resistance factor. The dfrF gene caused resistance in 4 out of 5 trimethoprim-resistant isolates from the German collection. An amino acid substitution in the intrinsic dihydrofolate reductase known from trimethoprim-resistant Streptococcus pneumoniae conferred resistance to S. pyogenes isolates of emm type 102.2, which lacked other aforementioned dfr genes. Trimethoprim may be more useful in treatment of S. pyogenes infections than previously thought. However, the factors described herein may lead to the rapid development and spread of resistance of S. pyogenes to this antibiotic agent. PMID:24492367

  3. Delineation of Streptococcus dysgalactiae, Its Subspecies, and Its Clinical and Phylogenetic Relationship to Streptococcus pyogenes

    PubMed Central

    Jensen, Anders

    2012-01-01

    The taxonomic status and structure of Streptococcus dysgalactiae have been the object of much confusion. Bacteria belonging to this species are usually referred to as Lancefield group C or group G streptococci in clinical settings in spite of the fact that these terms lack precision and prevent recognition of the exact clinical relevance of these bacteria. The purpose of this study was to develop an improved basis for delineation and identification of the individual species of the pyogenic group of streptococci in the clinical microbiology laboratory, with a special focus on S. dysgalactiae. We critically reexamined the genetic relationships of the species S. dysgalactiae, Streptococcus pyogenes, Streptococcus canis, and Streptococcus equi, which may share Lancefield group antigens, by phylogenetic reconstruction based on multilocus sequence analysis (MLSA) and 16S rRNA gene sequences and by emm typing combined with phenotypic characterization. Analysis of concatenated sequences of seven genes previously used for examination of viridans streptococci distinguished robust and coherent clusters. S. dysgalactiae consists of two separate clusters consistent with the two recognized subspecies dysgalactiae and equisimilis. Both taxa share alleles with S. pyogenes in several housekeeping genes, which invalidates identification based on single-locus sequencing. S. dysgalactiae, S. canis, and S. pyogenes constitute a closely related branch within the genus Streptococcus indicative of recent descent from a common ancestor, while S. equi is highly divergent from other species of the pyogenic group streptococci. The results provide an improved basis for identification of clinically important pyogenic group streptococci and explain the overlapping spectrum of infections caused by the species associated with humans. PMID:22075580

  4. Epidemiology of severe Streptococcus pyogenes disease in Europe.

    PubMed

    Lamagni, Theresa L; Darenberg, Jessica; Luca-Harari, Bogdan; Siljander, Tuula; Efstratiou, Androulla; Henriques-Normark, Birgitta; Vuopio-Varkila, Jaana; Bouvet, Anne; Creti, Roberta; Ekelund, Kim; Koliou, Maria; Reinert, Ralf René; Stathi, Angeliki; Strakova, Lenka; Ungureanu, Vasilica; Schalén, Claes; Jasir, Aftab

    2008-07-01

    The past 2 decades have brought worrying increases in severe Streptococcus pyogenes diseases globally. To investigate and compare the epidemiological patterns of these diseases within Europe, data were collected through a European Union FP-5-funded program (Strep-EURO). Prospective population-based surveillance of severe S. pyogenes infection diagnosed during 2003 and 2004 was undertaken in 11 countries across Europe (Cyprus, the Czech Republic, Denmark, Finland, France, Germany, Greece, Italy, Romania, Sweden, and the United Kingdom) using a standardized case definition. A total of 5,522 cases were identified across the 11 countries during this period. Rates of reported infection varied, reaching 3/100,000 population in the northern European countries. Seasonal patterns of infection showed remarkable congruence between countries. The risk of infection was highest among the elderly, and rates were higher in males than in females in most countries. Skin lesions/wounds were the most common predisposing factor, reported in 25% of cases; 21% had no predisposing factors reported. Skin and soft tissue were the most common foci of infection, with 32% of patients having cellulitis and 8% necrotizing fasciitis. The overall 7-day case fatality rate was 19%; it was 44% among patients who developed streptococcal toxic shock syndrome. The findings from Strep-EURO confirm a high incidence of severe S. pyogenes disease in Europe. Furthermore, these results have identified targets for public health intervention, as well as raising awareness of severe S. pyogenes disease across Europe.

  5. Pleiotropic virulence factor - Streptococcus pyogenes fibronectin-binding proteins.

    PubMed

    Yamaguchi, Masaya; Terao, Yutaka; Kawabata, Shigetada

    2013-04-01

    Streptococcus pyogenes causes a broad spectrum of infectious diseases, including pharyngitis, skin infections and invasive necrotizing fasciitis. The initial phase of infection involves colonization, followed by intimate contact with the host cells, thus promoting bacterial uptake by them. S. pyogenes recognizes fibronectin (Fn) through its own Fn-binding proteins to obtain access to epithelial and endothelial cells in host tissue. Fn-binding proteins bind to Fn to form a bridge to α5 β1 -integrins, which leads to rearrangement of cytoskeletal actin in host cells and uptake of invading S. pyogenes. Recently, several structural analyses of the invasion mechanism showed molecular interactions by which Fn converts from a compact plasma protein to a fibrillar component of the extracellular matrix. After colonization, S. pyogenes must evade the host innate immune system to spread into blood vessels and deeper organs. Some Fn-binding proteins contribute to evasion of host innate immunity, such as the complement system and phagocytosis. In addition, Fn-binding proteins have received focus as non-M protein vaccine candidates, because of their localization and conservation among different M serotypes.Here, we review the roles of Fn-binding proteins in the pathogenesis and speculate regarding possible vaccine antigen candidates. © 2012 Blackwell Publishing Ltd.

  6. Periorbital Necrotizing Fasciitis Secondary to Candida parapsilosis and Streptococcus pyogenes.

    PubMed

    Zhang, Matthew; Chelnis, James; Mawn, Louise A

    Necrotizing fasciitis is most often caused by either polymicrobial bacterial infections or by Gram-positive organisms, such as Streptococcus or Staphylococcus; however, rare cases of fungal necrotizing fasciitis have been reported. Candida parapsilosis is an emerging fungal pathogen. This fungus grows in either a yeast or pseudohyphal form. C. parapsilosis has been reported to cause keratitis, intraocular infection, and seeding of frontalis slings. C. parapsilosis is a commensal of human skin and can be acquired by nosocomial spread. Necrotizing fasciitis due to Candida has rarely been reported, but to date C. parapsilosis has not been identified as the causative organism in necrotizing fasciitis. This is the first documented case of human periocular soft tissue infection by C. parapsilosis, and also the first report providing evidence of mycotic infection in a necrotizing fasciitis concurrently infected by Streptococcus pyogenes.

  7. Cationic antimicrobial peptides disrupt the Streptococcus pyogenes ExPortal.

    PubMed

    Vega, Luis Alberto; Caparon, Michael G

    2012-09-01

    Although they possess a well-characterized ability to porate the bacterial membrane, emerging research suggests that cationic antimicrobial peptides (CAPs) can influence pathogen behaviour at levels that are sublethal. In this study, we investigated the interaction of polymyxin B and human neutrophil peptide (HNP-1) with the human pathogen Streptococcus pyogenes. At sublethal concentrations, these CAPs preferentially targeted the ExPortal, a unique microdomain of the S. pyogenes membrane, specialized for protein secretion and processing. A consequence of this interaction was the disruption of ExPortal organization and a redistribution of ExPortal components into the peripheral membrane. Redistribution was associated with inhibition of secretion of certain toxins, including the SpeB cysteine protease and the streptolysin O (SLO) cytolysin, but not SIC, a protein that protects S. pyogenes from CAPs. These data suggest a novel function for CAPs in targeting the ExPortal and interfering with secretion of factors required for infection and survival. This mechanism may prove valuable for the design of new types of antimicrobial agents to combat the emergence of antibiotic-resistant pathogens.

  8. Mechanisms of resistance for Streptococcus pyogenes in northern Utah.

    PubMed

    Rowe, Ryan A; Stephenson, Ryan M; East, Destry L; Wright, Scott

    2009-01-01

    The purpose of this study was to (1) determine the rates of penicillin and erythromycin resistance among Streptococcus pyogenes isolates in northern Utah, and (2) determine the genotype of the erythromycin resistant strains, thereby providing information regarding the mechanism of the resistance. Seven hundred thirty-nine isolates of S. pyogenes were identified on 5% Sheep Blood Agar. Susceptibility to erythromycin and penicillin was performed using Muller-Hinton blood agar. All isolates resistant to erythromycin were then genotyped using PCR primers specific to one of the following: mefA gene, indicating the mechanism of resistance was an efflux pump; ermA gene, in which the mechanism was inducible methylation of the ribosomes; and ermB indicating constitutive methylation of the ribosomes. This study was conducted at Weber State University, in the Department of Clinical Laboratory Sciences. Samples were collected from 9 clinics ranging from North Ogden to Taylorsville, Utah. All samples were previously tested positive for S. pyogenes by the clinic from where the samples were collected. Of the 739 S. pyogenes isolates tested, 2.4% were resistant to erythromycin with no resistance observed to penicillin. Of the strains that displayed some degree of resistance, the gene frequencies observed were as follows: 48.1% mefA, 26.0% ermA, 3.7% ermB, and 22.2% multiple genes. The most common genotype was mefA, indicating that the efflux pump (M phenotype) is the most common mechanism in the surveyed area, followed by ermA, which produces the inducible methylating enzyme. A significant number of isolates was also observed to express both the efflux pump and the constitutive methylating enzyme.

  9. Metabolic effects of static magnetic fields on Streptococcus pyogenes.

    PubMed

    Morrow, A C; Dunstan, R H; King, B V; Roberts, T K

    2007-09-01

    This study aimed to develop a simple experimental system utilising bacterial cells to investigate the dose responses resulting from exposures to static magnetic flux densities ranging from 0.05 to 0.5 T on viability, bacterial metabolism and levels of DNA damage in Streptococcus pyogenes. Exposure of S. pyogenes to a field of 0.3 T at 24 degrees C under anaerobic conditions resulted in a significant (P < 0.05) decrease in growth rate, with an increased mean generation time of 199 +/- 6 min compared to the control cells at 165 +/- 6 min (P < 0.05). Conversely, exposure to magnetic fields of 0.5 T significantly accelerated the growth rate at 24 degrees C compared to control cells, with a decreased mean generation time of 147 +/- 4 min (P < 0.05). The patterns of metabolite release from cells incubated in phosphate buffered saline (PBS) at 24 degrees C and exposed to different magnetic flux densities (0.05-0.5 T) were significantly (P < 0.05) altered, compared to non-exposed controls. Concentrations of metabolites, with the exception of aspartic acid (r = 0.44), were not linearly correlated with magnetic flux density, with all other r < 0.20. Instead, "window" effects were observed, with 0.25-0.3 T eliciting the maximal release of the majority of metabolites, suggesting that magnetic fields of these strengths had significant impacts on metabolic homeostasis in S. pyogenes. The exposure of cells to 0.3 T was also found to significantly reduce the yield of 8-hydroxyguanine in extracted DNA compared to controls, suggesting some possible anti-oxidant protection to S. pyogenes at this field strength.

  10. Severe Streptococcus pyogenes Infections, United Kingdom, 2003–2004

    PubMed Central

    Neal, Shona; Keshishian, Catherine; Alhaddad, Neelam; George, Robert; Duckworth, Georgia; Vuopio-Varkila, Jaana; Efstratiou, Androulla

    2008-01-01

    As part of a Europe-wide initiative to explore current epidemiologic patterns of severe disease caused by Streptococcus pyogenes, the United Kingdom undertook enhanced population-based surveillance during 2003–2004. A total of 3,775 confirmed cases of severe S. pyogenes infection were identified over 2 years, 3.33/100,000 population, substantially more than previously estimated. Skin/soft tissue infections were the most common manifestation (42%), followed by respiratory tract infections (17%). Injection drug use was identified as a risk factor for 20% of case-patients. One in 5 infected case-patients died within 7 days of diagnosis; the highest mortality rate was for cases of necrotizing fasciitis (34%). Nonsteroidal antiinflammatory drugs, alcoholism, young age, and infection with emm/M3 types were independently associated with increased risk for streptococcal toxic shock syndrome. Understanding the pattern of these diseases and predictors of poor patient outcome will help with identification and assessment of the potential effect of targeted interventions. PMID:18258111

  11. Severe Streptococcus pyogenes infections, United Kingdom, 2003-2004.

    PubMed

    Lamagni, Theresa L; Neal, Shona; Keshishian, Catherine; Alhaddad, Neelam; George, Robert; Duckworth, Georgia; Vuopio-Varkila, Jaana; Efstratiou, Androulla

    2008-02-01

    As part of a Europe-wide initiative to explore current epidemiologic patterns of severe disease caused by Streptococcus pyogenes, the United Kingdom undertook enhanced population-based surveillance during 2003-2004. A total of 3,775 confirmed cases of severe S. pyogenes infection were identified over 2 years, 3.33/100,000 population, substantially more than previously estimated. Skin/soft tissue infections were the most common manifestation (42%), followed by respiratory tract infections (17%). Injection drug use was identified as a risk factor for 20% of case-patients. One in 5 infected case-patients died within 7 days of diagnosis; the highest mortality rate was for cases of necrotizing fasciitis (34%). Nonsteroidal antiinflammatory drugs, alcoholism, young age, and infection with emm/M3 types were independently associated with increased risk for streptococcal toxic shock syndrome. Understanding the pattern of these diseases and predictors of poor patient outcome will help with identification and assessment of the potential effect of targeted interventions.

  12. A novel adaptation of aldolase regulates virulence in Streptococcus pyogenes

    PubMed Central

    Loughman, Jennifer A; Caparon, Michael G

    2006-01-01

    Regulation of virulence factor expression is critical for pathogenic microorganisms that must sense and adapt to a dynamic host environment; yet, the signal transduction pathways that enable this process are generally poorly understood. Here, we identify LacD.1 as a global regulator of virulence factor expression in the versatile human pathogen, Streptococcus pyogenes. LacD.1 is derived from a class I tagatose-1,6-bisphosphate aldolase homologous to those involved in lactose and galactose metabolism in related prokaryotes. However, regulation of transcription by LacD.1 is not dependent on this enzymatic activity or the canonical catabolite repression pathway, but likely does require substrate recognition. Our results suggest that LacD.1 has been adapted as a metabolic sensor, and raise the possibility that regulation of gene expression by metabolic enzymes may be a novel mechanism by which Gram-positive bacteria, including S. pyogenes, coordinate multiple environmental cues, allowing essential transcription programs to be coupled with perceived nutritional status. PMID:17066081

  13. Macrolide-Resistant Streptococcus pyogenes in Norway: Population Structure and Resistance Determinants

    PubMed Central

    Littauer, P.; Caugant, D. A.; Sangvik, M.; Høiby, E. A.; Sundsfjord, A.; Simonsen, G. S.

    2006-01-01

    A 2.7% prevalence of macrolide resistance in 1,657 Norwegian clinical Streptococcus pyogenes isolates was primarily due to erm(TR) (59%) and mef(A) (20%). Four clonal complexes comprised 75% of the strains. Macrolide resistance in S. pyogenes in Norway is imported as resistant strains or locally selected in internationally disseminated susceptible clones. PMID:16641473

  14. The streptococcal inhibitor of complement (SIC) protects Streptococcus pyogenes from bacteriocin-like inhibitory substance (BLIS) from Streptococcus salivarius.

    PubMed

    Minami, Masaaki; Ohmori, Daisuke; Tatsuno, Ichiro; Isaka, Masanori; Kawamura, Yoshiaki; Ohta, Michio; Hasegawa, Tadao

    2009-09-01

    Streptococcus salivarius inhibits the growth of Streptococcus pyogenes in vitro. Streptococcus pyogenes has various virulence factors, including the streptococcus inhibitor of complement (SIC). Although SIC inhibits the activity of the peptides LL-37 and NAP1, the relationship between SIC and the bacteriocin-like inhibitory substance (BLIS) has not been elucidated. Here, we evaluated whether S. salivarius BLIS affects S. pyogenes SIC. We created three deltasic mutant strains from three S. pyogenes strains and performed deferred antagonism assays. The test strains were BLIS-positive S. salivarius JCM5707 and BLIS-negative S. salivarius NCU12. Deferred antagonism assays with JCM5707 showed that the inhibitory zones in the three deltasic mutant strains were wider than those in the three wild-type strains. Streptococcus pyogenes was cultured in BLIS-containing broth and the change in SIC in the supernatant was assessed by two-dimensional gel electrophoresis (2-DE). The 2-DE analysis of S. pyogenes exoproteins with the JCM5707 supernatant showed reduced SIC compared with those without the JCM5707 supernatant. Changes in sic mRNA levels affected by S. salivarius BLIS were evaluated by a reverse transcriptase-PCR. The sic mRNA level was affected more by the BLIS-positive S. salivarius than by the BLIS-negative strain. Our result indicates that SIC plays a role in the inhibition of S. salivarius BLIS.

  15. [Streptococcus pyogenes and the brain: living with the enemy].

    PubMed

    Dale, R C

    Streptococcus pyogenes (or group A beta hemolytic streptococcus) is a pathogenic bacterium that can give rise to a range of invasive and autoimmune diseases, although it is more widely known as the cause of tonsillitis. It is particularly interesting to note that this germ only causes disease in humans. For many years it has been acknowledged that it can cause an autoimmune brain disease (Sydenham s chorea). Yet, the spectrum of post streptococcal brain disorders has recently been extended to include other movement disorders such as tics or dystonia. A number of systematic psychiatric studies have shown that certain emotional disorders generally accompany the movement disorder (particularly, obsessive compulsive disorder). The proposed pathogenetic mechanism is that of a neuronal dysfunction in which antibodies play a mediating role. The antibodies that are produced after the streptococcal infection cross react with neuronal proteins, and more especially so in individuals with a propensity. This represents a possible model of immunological mimicry and its potential importance with respect to certain idiopathic disorders such as Tourette syndrome and obsessive compulsive disorder.

  16. Molecular Epidemiology of sil Locus in Clinical Streptococcus pyogenes Strains

    PubMed Central

    Plainvert, Céline; Dinis, Márcia; Ravins, Miriam; Hanski, Emanuel; Touak, Gérald; Dmytruk, Nicolas; Fouet, Agnès

    2014-01-01

    Streptococcus pyogenes (group A Streptococcus [GAS]) causes a wide variety of diseases, ranging from mild noninvasive to severe invasive infections. Mutations in regulatory components have been implicated in the switch from colonization to invasive phenotypes. The inactivation of the sil locus, composed of six genes encoding a quorum-sensing complex, gives rise to a highly invasive strain. However, studies conducted on limited collections of GAS strains suggested that sil prevalence is around 15%; furthermore, whereas a correlation between the presence of sil and the genetic background was suggested, no link between the presence of a functional sil locus and the invasive status was assessed. We established a collection of 637 nonredundant strains covering all emm genotypes present in France and of known clinical history; 68%, 22%, and 10% were from invasive infections, noninvasive infections, and asymptomatic carriage, respectively. Among the 637 strains, 206 were sil positive. The prevalence of the sil locus varied according to the emm genotype, being present in >85% of the emm4, emm18, emm32, emm60, emm87, and emm90 strains and absent from all emm1, emm28, and emm89 strains. A random selection based on 2009 French epidemiological data indicated that 16% of GAS strains are sil positive. Moreover, due to mutations leading to truncated proteins, only 9% of GAS strains harbor a predicted functional sil system. No correlation was observed between the presence or absence of a functional sil locus and the strain invasiveness status. PMID:24671796

  17. Nonhemolytic Streptococcus pyogenes Isolates That Lack Large Regions of the sag Operon Mediating Streptolysin S Production▿

    PubMed Central

    Yoshino, Miho; Murayama, Somay Y.; Sunaoshi, Katsuhiko; Wajima, Takeaki; Takahashi, Miki; Masaki, Junko; Kurokawa, Iku; Ubukata, Kimiko

    2010-01-01

    Among nonhemolytic Streptococcus pyogenes (group A streptococcus) strains (n = 9) isolated from patients with pharyngitis or acute otitis media, we identified three deletions in the region from the epf gene, encoding the extracellular matrix binding protein, to the sag operon, mediating streptolysin S production. PMID:20018818

  18. Antibiotic Selection Pressure and Resistance in Streptococcus pneumoniae and Streptococcus pyogenes

    PubMed Central

    Albrich, Werner C.; Monnet, Dominique L.

    2004-01-01

    We correlated outpatient antibiotic use with prevalence of penicillin-nonsusceptible Streptococcus pneumoniae (PNSP), macrolide-resistant S. pneumoniae (MRSP), and macrolide-resistant S. pyogenes (MRGAS) in 20 countries. Total antibiotic use was correlated with PNSP (r = 0.75; p < 0.001), as was macrolide use with MRSP (r = 0.88; p < 0.001) and MRGAS (r = 0.71; p = 0.004). Streptococcal resistance is directly associated with antibiotic selection pressure on a national level. PMID:15109426

  19. [Clustered cases of intrafamily invasive Streptococcus pyogenes infection (or group A streptococcus)].

    PubMed

    Caillet-Gossot, S; Rousset-Rouviere, C; Arlaud, K; Dubus, J-C; Bosdure, E

    2011-12-01

    Streptococcus pyogenes or group A streptococcus (GAS) is responsible for serious invasive infections with a risk of secondary infection in patients with more contact than in the general population. Regardless of clustering, few intrafamilial invasive infections have been reported despite a recent increase in the incidence of invasive GAS disease. We report the cases of two brothers, one a boy of 8.5 years with toxic shock syndrome with no bacteria identified and the second, 1 week later, his 14.5-year-old brother in hospital for sepsis due to GAS. The occurrence of a confirmed case of invasive GAS and a probable case within such a short period met the definition of clustered cases. Both brothers showed no risk factors for invasive disease and no gateway including skin was found. Antibiotic therapy was initiated in the family as recommended by the French Higher Council of Public Hygiene. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

  20. [Streptococcus pyogenes--much more than the aetiological agent of scarlet fever].

    PubMed

    Stock, Ingo

    2009-11-01

    The grampositive bacterium S. pyogenes (beta-haemolytic group A Streptococcus) is a natural colonizer of the human oropharynx mucous membrane and one of the most common agents of infectious diseases in humans. S. pyogenes causes the widest range of disease in humans among all bacterial pathogens. It is responsible for various skin infections such as impetigo contagiosa and erysipelas, and localized mucous membrane infections of the oropharynx (e. g. tonsillitis and pharyngitis). Betahaemolytic group A Streptococcus causes also invasive diseases such as sepses including puerperal sepsis. Additionally, S. pyogenes induces toxin-mediated syndromes, i. e. scarlet fever, streptococcal toxic shock syndrome (STSS) and necrotizing fasciitis (NF). STSS and NF are severe, frequently fatal diseases that have emerged in Europe and Northern America during the last two decades. Finally, some immunpathological diseases such as acute rheumatic fever and glomerulonephritis also result from S. pyogenes infections. Most scientists recommend penicillins (benzylpenicillin, phenoxymethylpenicllin) as drugs of first choice for treatment of Streptococcus tonsillopharyngitis and scarlet fever. Erysipelas and some other skin infections should be treated with benzylpenicillin. Intensive care measurements are needed for treatment of severe toxin-mediated S. pyogenes diseases. These measurements include the elimination of internal bacterial foci, concomitant application of clindamycin and benzylpenicillin and suitable treatment of shock symptoms. Management of immunpathological diseases requires antiphlogistical therapy. Because of the wide distribution of S. pyogenes in the general population and the lack of an effective vaccine, possibilities for prevention allowing a suitable protection for diseases due to S. pyogenes are very limited.

  1. Streptococcus pyogenes biofilms-formation, biology, and clinical relevance.

    PubMed

    Fiedler, Tomas; Köller, Thomas; Kreikemeyer, Bernd

    2015-01-01

    Streptococcus pyogenes (group A streptococci, GAS) is an exclusive human bacterial pathogen. The virulence potential of this species is tremendous. Interactions with humans range from asymptomatic carriage over mild and superficial infections of skin and mucosal membranes up to systemic purulent toxic-invasive disease manifestations. Particularly the latter are a severe threat for predisposed patients and lead to significant death tolls worldwide. This places GAS among the most important Gram-positive bacterial pathogens. Many recent reviews have highlighted the GAS repertoire of virulence factors, regulators and regulatory circuits/networks that enable GAS to colonize the host and to deal with all levels of the host immune defense. This covers in vitro and in vivo studies, including animal infection studies based on mice and more relevant, macaque monkeys. It is now appreciated that GAS, like many other bacterial species, do not necessarily exclusively live in a planktonic lifestyle. GAS is capable of microcolony and biofilm formation on host cells and tissues. We are now beginning to understand that this feature significantly contributes to GAS pathogenesis. In this review we will discuss the current knowledge on GAS biofilm formation, the biofilm-phenotype associated virulence factors, regulatory aspects of biofilm formation, the clinical relevance, and finally contemporary treatment regimens and future treatment options.

  2. Streptococcus pyogenes bacteraemia, emm types and superantigen profiles.

    PubMed

    Rantala, S; Vähäkuopus, S; Siljander, T; Vuopio, J; Huhtala, H; Vuento, R; Syrjänen, J

    2012-05-01

    The aim of this study was to investigate the emm types and superantigen profiles of bacteraemic group A streptococcal (GAS; Streptococcus pyogenes) isolates and to detect possible associations between the molecular characteristics of isolates and the clinical presentations of disease. In this population-based study, 87 bacteraemic GAS isolates from adult patients in Pirkanmaa Health District (HD), Finland, during the period 1995-2004 were emm typed and genotyped for superantigen (SAg) profiles. The epidemiological and clinical data of the patients were analysed with the microbiological characterisation data. Among the 87 isolates, 18 different emm types were found. emm1, emm28 and emm81 were the three most common types, covering 52% of isolates. The prevalence of specific emm types showed high variability during the 10-year study period. We could not find any association between the emm type and clinical features of bacteraemic infection, such as underlying diseases, disease manifestations or case fatality. Of nine superantigen genes examined, speA and speC were identified in 20 and 30% of the strains, respectively. No association was found between disease manifestation and the presence of single superantigen genes. The 26-valent GAS vaccine would have covered only 62% of isolates causing invasive disease in Pirkanmaa HD during the study period.

  3. Streptococcus pyogenes biofilms—formation, biology, and clinical relevance

    PubMed Central

    Fiedler, Tomas; Köller, Thomas; Kreikemeyer, Bernd

    2015-01-01

    Streptococcus pyogenes (group A streptococci, GAS) is an exclusive human bacterial pathogen. The virulence potential of this species is tremendous. Interactions with humans range from asymptomatic carriage over mild and superficial infections of skin and mucosal membranes up to systemic purulent toxic-invasive disease manifestations. Particularly the latter are a severe threat for predisposed patients and lead to significant death tolls worldwide. This places GAS among the most important Gram-positive bacterial pathogens. Many recent reviews have highlighted the GAS repertoire of virulence factors, regulators and regulatory circuits/networks that enable GAS to colonize the host and to deal with all levels of the host immune defense. This covers in vitro and in vivo studies, including animal infection studies based on mice and more relevant, macaque monkeys. It is now appreciated that GAS, like many other bacterial species, do not necessarily exclusively live in a planktonic lifestyle. GAS is capable of microcolony and biofilm formation on host cells and tissues. We are now beginning to understand that this feature significantly contributes to GAS pathogenesis. In this review we will discuss the current knowledge on GAS biofilm formation, the biofilm-phenotype associated virulence factors, regulatory aspects of biofilm formation, the clinical relevance, and finally contemporary treatment regimens and future treatment options. PMID:25717441

  4. Comparative functional analysis of the lac operons in Streptococcus pyogenes.

    PubMed

    Loughman, Jennifer A; Caparon, Michael G

    2007-04-01

    Having no known environmental reservoir, Streptococcus pyogenes, a bacterium responsible for a wider variety of human diseases than any other bacterial species, must rely on its host for metabolic substrates. Although a streptococcal aldolase, LacD.1, has been adapted to virulence gene regulation, both LacD.1 and a paralogous protein, LacD.2, are predicted to function in the tagatose 6-phosphate pathway for lactose and galactose utilization. In order to gain insight into the mechanism of the LacD.1 regulatory pathway and the role of genome context in the emergence of LacD.1's novel regulatory functions, we compared the function and regulation of the Lac.1 and Lac.2 loci. The Lac.1 operon is not inducible, and regulation by LacD.1 is independent of a functional tagatose 6-phosphate pathway and enhanced by the conserved truncation of upstream Lac.1 genes. In contrast, Lac.2 expression is sensitive to environmental carbohydrates, and LacD.2, not LacD.1, contributes to growth on galactose. Thus, we conclude that the Lac.1 locus has been specialized to participate in regulation, leaving efficient utilization of carbohydrate sources to the Lac.2 locus. The adaptation of LacD for transcription regulation may be an underappreciated strategy among prokaryotes, as homologues of this multifaceted enzyme are present in a broad range of species.

  5. Vaccination against the M protein of Streptococcus pyogenes prevents death after influenza virus: S. pyogenes super-infection.

    PubMed

    Klonoski, Joshua M; Hurtig, Heather R; Juber, Brian A; Schuneman, Margaret J; Bickett, Thomas E; Svendsen, Joshua M; Burum, Brandon; Penfound, Thomas A; Sereda, Grigoriy; Dale, James B; Chaussee, Michael S; Huber, Victor C

    2014-09-08

    Influenza virus infections are associated with a significant number of illnesses and deaths on an annual basis. Many of the deaths are due to complications from secondary bacterial invaders, including Streptococcus pneumoniae, Staphylococcus aureus, Haemophilus influenzae, and Streptococcus pyogenes. The β-hemolytic bacteria S. pyogenes colonizes both skin and respiratory surfaces, and frequently presents clinically as strep throat or impetigo. However, when these bacteria gain access to normally sterile sites, they can cause deadly diseases including sepsis, necrotizing fasciitis, and pneumonia. We previously developed a model of influenza virus:S. pyogenes super-infection, which we used to demonstrate that vaccination against influenza virus can limit deaths associated with a secondary bacterial infection, but this protection was not complete. In the current study, we evaluated the efficacy of a vaccine that targets the M protein of S. pyogenes to determine whether immunity toward the bacteria alone would allow the host to survive an influenza virus:S. pyogenes super-infection. Our data demonstrate that vaccination against the M protein induces IgG antibodies, in particular those of the IgG1 and IgG2a isotypes, and that these antibodies can interact with macrophages. Ultimately, this vaccine-induced immunity eliminated death within our influenza virus:S. pyogenes super-infection model, despite the fact that all M protein-vaccinated mice showed signs of illness following influenza virus inoculation. These findings identify immunity against bacteria as an important component of protection against influenza virus:bacteria super-infection. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Colonisation with Staphylococcus aureus and Streptococcus pyogenes in New Zealand preschool children.

    PubMed

    Berry, Sarah; Morton, Susan; Atatoa Carr, Polly; Marks, Emma; Ritchie, Stephen; Upton, Arlo; Williamson, Debbie; Grant, Cameron

    2015-03-13

    To describe colonisation patterns of Staphylococcus aureus (S. aureus) and Streptococcus pyogenes (S. pyogenes) among pre-school children in New Zealand. Anterior nasal, oropharyngeal, and antecubital fossa swabs were collected from a diverse sample of 139 New Zealand children aged 4 years. Swabs were cultured for S. aureus and S. pyogenes. S. aureus isolates were tested for antibiotic susceptibility. S. aureus colonisation was more prevalent than S. pyogenes colonisation; 54% of the children were colonised with S. aureus whereas only 16% were colonised with S. pyogenes, at one or more sampling sites (P<0.0001). S. aureus was present in a larger proportion of swabs obtained from the anterior nasal (39%, P<0.0001) or oropharynx (32%, P=0.0002) than from the antecubital fossa (14%). S. pyogenes was present in a larger proportion of swabs obtained from the oropharynx (16%) than either the anterior nasal (4%, P=0.001) or the antecubital fossa (2%, P<0.0001). S. aureus and S. pyogenes are prevalent at superficial sites in preschool children in NZ, with S. aureus colonisation more prevalent than S. pyogenes colonisation. Colonisation frequency varies by site for both pathogens; S. aureus is more prevalent in the anterior nares and oropharynx while S. pyogenes is more prevalent in the oropharynx.

  7. Bacterial Pneumonia Caused by Streptococcus pyogenes Infection: A Case Report and Review of the Literature

    PubMed Central

    Akuzawa, Nobuhiro; Kurabayashi, Masahiko

    2016-01-01

    A 78-year-old Japanese man was admitted to our hospital because of fever lasting for 4 days. His white blood cell count and C-reactive protein level were elevated and computed tomography of the chest showed bronchopneumonia in the right upper lobe of the lung. Streptococcus pyogenes was detected from sputum and blood culture samples on admission and administration of ampicillin/sulbactam was effective. Although our patient’s clinical course was good, S. pyogenes pneumonia commonly shows a high rate of fatality and septicemia, and may affect a previously healthy population. Physicians should be aware of pernicious characteristics of S. pyogenes pneumonia. PMID:27738486

  8. [Streptococcus pyogenes infection in paediatrics: from pharyngotonsillitis to invasive infections].

    PubMed

    Espadas Maciá, David; Flor Macián, Eva María; Borrás, Rafael; Poujois Gisbert, Sandrine; Muñoz Bonet, Juan Ignacio

    2017-03-31

    Streptococcus pyogenes or Group A Streptococci (GAS) cause many infections in infancy. Changes in its epidemiology have been described in recent years, including an increase in invasive infections (iGAS). A retrospective-descriptive study was conducted on children less than 15 years old, with GAS infections, in particular iGAS, and their complications from February 2004-April 2014. A total of 2,192 positive cultures were obtained of which 92.7% were pharyngeal cultures. Twenty-nine patients were admitted to hospital: 4 with suppurative complications, 7 post-infective, 14 iGAS, and 4 probable iGAS cases. There were no differences in the frequency of GAS isolations/year. Non-invasive isolates were more frequent in winter and spring (P<.001), and 68.3% were in patients younger than 5 years. The incidence of iGAS was 2.1/100,000 children/year. There was no seasonality, and it was more frequent in younger children (P=.039). The most common diagnosis was pneumonia (6/14). Eight patients required intensive care. They were treated empirically with second or third-generation cephalosporin or with intravenous penicillin, and pneumonia required longer treatment times (P=.016). All GAS isolates were sensitive to penicillin, and 10.6% were resistant to erythromycin. The time spent in hospital was longer for iGAS than other cases (P=.028). No patients died. Pharyngotonsillitis caused by GAS is common in childhood, and its incidence is increasing in children younger than 5 years. At the moment, post-infectious complications are rare. Invasive infections are the most severe forms of presentation, and are more common in younger children. Copyright © 2017. Publicado por Elsevier España, S.L.U.

  9. Invasive disease by Streptococcus pyogenes: patients hospitalized for 6 years.

    PubMed

    Arias-Constantí, Vanessa; Trenchs-Sainz de la Maza, Victoria; Sanz-Marcos, Nuria Elvira; Guitart-Pardellans, Carmina; Gené-Giralt, Amadeu; Luaces-Cubells, Carles

    2017-07-10

    The last years an increase of severe cases of invasive disease (ID) due to Streptococcus pyogenes or streptococcus b-hemolytic group A (SGA) had been detected. The aim of this study was to analyze the epidemiology and the clinical features of ID due to SGA in a tertiary Pediatric Hospital. Retrospective study in a Pediatric hospital, of all in-patients with final diagnosis of ID due to SGA during 6 years (2009-2014). To consider ID, SGA had to be isolated in sterile samples; in patients with fascitis necroticans in skin samples or in any sample in patients with the diagnostic of Streptococcal Toxic Shock Syndrome (STSS). The SSTS was defined as hypotension and at least 2 of these criteria: renal failure, hepatic failure, acute respiratory distress, tissue necrosis or desquamative erythematous rash. Demographic data, type of infection, risk factors, clinical presentation, analytical data at admission, treatment, need for admission to a pediatric intensive care unit, microbiological data, hospital stay and evolution were collected. Fifty-two (52) cases were included (12/10,000 of all inpatients); 3 years-old was the medium age (p25-75: 1.4-6.9 years); 28 (53.8%) were boys. Fourteen patients (26.9%) had risk factors. Fever was the major symptom (51 patients, 98.1%). The skin lesions were the most frequent clinical manifestations found (21; 40.4%). In 50 (96%) cases, SGA was isolated in at least one sterile sample. Skin and soft tissue infections were diagnosed in 14 patients (26.9%), 14 (26.9%) pneumonias, 12 (23.1%) bones and joints infections, 10 (19.2%) SSTS, 6 (11.5%) occult bacteremia, 4 (7.7%) meningitis and 2 (3.8%) sepsis. Surgery was required in 18 cases (34.6%) and 17 patients (32.7%) needed intensive care. The medium hospital stay was 9.5 days (p25-75: 8-15 days). Three patients presented sequels and one patient died. The ID due to SGA was a rare but serious reason for hospital admission. Skin and soft tissue infections, and pleuroneumonia were the most

  10. Protective effect of hainosankyuto, a traditional Japanese medicine, on Streptococcus pyogenes infection in murine model.

    PubMed

    Minami, Masaaki; Ichikawa, Mariko; Hata, Nanako; Hasegawa, Tadao

    2011-01-01

    Streptococcus pyogenes (S. pyogenes) causes various serious diseases including necrotizing fasciitis and streptococcal toxic shock syndrome. One serious problem observed recently with S. pyogenes therapy is attenuation of the antibiotic effect, especially penicillin treatment failure and macrolide resistance. Hainosankyuto, a traditional Japanese medicine based on ancient Chinese medicine, has been used for treatment of infectious purulent diseases in Japan. In this study, we investigated the protective and therapeutic efficacy of Hainosankyuto against S. pyogenes-skin infection. A broth microdilution method revealed that Hainosankyuto did not show a direct anti-bacterial effect against S. pyogenes. Force-feeding Hainosankyuto to infected mice for 4 consecutive days increased the survival rate and reduced the size of local skin lesions compared with mice fed PBS. Although we did not find the significant recovery of survival rate in Hainosankyuto administration only after S. pyogenes infection, the sizes of ulcer lesion were significant smaller after Hainosankyuto administration compared with mice fed PBS. No difference was observed in the anti-bacterial effect of Hainosankyuto between macrolide-susceptible and -resistant strains. Blood bactericidal assay showed that the survival rate of S. pyogenes using the blood from Hainosankyuto-treated mice was lower than that using the blood from untreated mice. We also found increased levels of IL-12, IFN-γ and a decreased level of TNF-α in the serum of S. pyogenes-infected mice treated with Hainosankyuto. Mouse peritoneal macrophage from Hainosankyuto-treated mice had significant phagocytic activity and increased mRNA levels of IL-12, IFN-γ and decreased mRNA level of TNF-α compared with control macrophage. Hainosankyuto increased survival rate after S. pyogenes infection and up-regulated both blood bactericidal activity and macrophage phagocytic activity through modulation of inflammatory cytokines. Our data also

  11. Incomplete Kawasaki disease associated with complicated Streptococcus pyogenes pneumonia: A case report.

    PubMed

    Leahy, Timothy Ronan; Cohen, Eyal; Allen, Upton D

    2012-01-01

    A three-year-old boy presented with community-acquired pneumonia complicated by empyema. Streptococcus pyogenes (group A streptococcus) was identified on culture of the pleural fluid. The patient improved with antibiotic therapy and drainage of the empyema. During his convalescence, the patient developed persistent fever, lethargy and anorexia. His inflammatory markers were elevated, and repeat cultures were negative. Although the patient had none of the classical mucocutaneous features of Kawasaki disease, an echocardiogram was performed, which revealed coronary artery dilation. The patient was diagnosed with incomplete Kawasaki disease and treated with intravenous immunoglobulin and high-dose acetylsalicylic acid. The fever subsided within 48 h. To the authors' knowledge, the present report is the first report of Kawasaki disease associated with complicated S pyogenes pneumonia. It emphasizes the importance of considering incomplete Kawasaki disease among children with persistent fever, the role of echocardiography in diagnosis, and the potential link between Kawasaki disease and superantigen-producing organisms such as S pyogenes.

  12. Recombination between Streptococcus suis ICESsu32457 and Streptococcus agalactiae ICESa2603 yields a hybrid ICE transferable to Streptococcus pyogenes.

    PubMed

    Marini, Emanuela; Palmieri, Claudio; Magi, Gloria; Facinelli, Bruna

    2015-07-09

    Integrative conjugative elements (ICEs) are mobile genetic elements that reside in the chromosome but retain the ability to undergo excision and to transfer by conjugation. Genes involved in drug resistance, virulence, or niche adaptation are often found among backbone genes as cargo DNA. We recently characterized in Streptococcus suis an ICE (ICESsu32457) carrying resistance genes [tet(O/W/32/O), tet(40), erm(B), aphA, and aadE] in the 15K unstable genetic element, which is flanked by two ∼1.3kb direct repeats. Remarkably, ∼1.3-kb sequences are conserved in ICESa2603 of Streptococcus agalactiae 2603V/R, which carry heavy metal resistance genes cadC/cadA and mer. In matings between S. suis 32457 (donor) and S. agalactiae 2603V/R (recipient), transconjugants were obtained. PCR experiments, PFGE, and sequence analysis of transconjugants demonstrated a tandem array between ICESsu32457 and ICESa2603. Matings between tandem array-containing S. agalactiae 2603V/R (donor) and Streptococcus pyogenes RF12 (recipient) yielded a single transconjugant containing a hybrid ICE, here named ICESa2603/ICESsu32457. The hybrid formed by recombination of the left ∼1.3-kb sequence of ICESsu32457 and the ∼1.3-kb sequence of ICESa2603. Interestingly, the hybrid ICE was transferable between S. pyogenes strains, thus demonstrating that it behaves as a conventional ICE. These findings suggest that both tandem arrays and hybrid ICEs may contribute to the evolution of antibiotic resistance in streptococci, creating novel mobile elements capable of disseminating new combinations of antibiotic resistance genes.

  13. Draft Genome Sequences of Streptococcus pyogenes Strains Associated with Throat and Skin Infections in Lebanon.

    PubMed

    Tokajian, Sima; Eisen, Jonathan A; Jospin, Guillaume; Coil, David A

    2014-05-15

    We present the draft genome sequences of nine clinical Streptococcus pyogenes isolates recovered from patients suffering from sore throat and skin infections. An average of 2,454,334 paired-end reads per sample were generated, which assembled into 21 to 198 contigs, with a G+C content of 38.4 to 38.5%. Copyright © 2014 Tokajian et al.

  14. Analysis of beta-hemolysis in human blood agars by Streptococcus pyogenes.

    PubMed

    Zomorodian, Kamiar; Rahimi, Mohammad Javad; Safaei, Akbar; Bazargani, Abdollah; Motamadi, Marjan; Kharazi, Mahboobeh; Mostaghni, Setareh; Pakshir, Keyvan; Ghaedi, Hamid; Afsarian, Mohammad Hossein

    2011-06-01

    The aim of the study was to assess the reliability of human blood agar media (HuBA) in identifying Streptococcus pyogenes by hemolysis analysis. We analyze several factors that might affect the accuracy of HuBA media for microbial analysis, including incubation time, blood group, Rh factor and presence of antistreptolysin-o.

  15. High prevalence of fluoroquinolone-nonsusceptible Streptococcus pyogenes emm12 in Taiwan.

    PubMed

    Lin, Jiun-Nong; Chang, Lin-Li; Lai, Chung-Hsu; Huang, Yi-Han; Chen, Wei-Fang; Yang, Chih-Hui; Hsu, Janine; Lin, Hsi-Hsun; Chen, Yen-Hsu

    2015-10-01

    Fluoroquinolone-nonsusceptible Streptococcus pyogenes has rapidly emerged in several countries. The aim of this study was to survey the epidemiology and molecular characteristics of fluoroquinolone-nonsusceptible S. pyogenes in Taiwan. A total of 350 consecutive S. pyogenes isolates were collected between January 2005 and December 2012, including 152 (43.4%) invasive and 198 (56.6%) noninvasive isolates. Thirty-nine isolates (11.1%) of S. pyogenes were nonsusceptible to fluoroquinolones, including one emm1/ST28, 4 emm4/ST39, 33 emm12/ST36, and 1 emm87/ST62. Of all the isolates, emm12 (50%) demonstrated the highest prevalence of fluoroquinolone nonsusceptibility. Alterations of Ser79Phe and Ala12Val in ParC were the most frequently mutations in fluoroquinolone-nonsusceptible S. pyogenes isolates. There were no amino acid substitutions in GyrB, and 1 emm87 isolate exhibited 3 nonsynonymous mutations in ParE. Our study reveals the emergence of fluoroquinolone-nonsusceptible S. pyogenes emm12/ST36 in Taiwan. Regular surveillance of fluoroquinolone susceptibility in S. pyogenes is suggested. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. CNS autoimmune disease after Streptococcus pyogenes infections: animal models, cellular mechanisms and genetic factors

    PubMed Central

    Cutforth, Tyler; DeMille, Mellissa MC; Agalliu, Ilir; Agalliu, Dritan

    2016-01-01

    Streptococcus pyogenes infections have been associated with two autoimmune diseases of the CNS: Sydenham’s chorea (SC) and Pediatric Autoimmune Neuropsychiatric Disorders Associated with Streptococcus infections (PANDAS). Despite the high frequency of pharyngeal streptococcus infections among children, only a small fraction develops SC or PANDAS. This suggests that several factors in combination are necessary to trigger autoimmune complications: specific S. pyogenes strains that induce a strong immune response toward the host nervous system; genetic susceptibility that predispose children toward an autoimmune response involving movement or tic symptoms; and multiple infections of the throat or tonsils that lead to a robust Th17 cellular and humoral immune response when untreated. In this review, we summarize the evidence for each factor and propose that all must be met for the requisite neurovascular pathology and behavioral deficits found in SC/PANDAS. PMID:27110222

  17. Chromosomal islands of Streptococcus pyogenes and related streptococci: molecular switches for survival and virulence.

    PubMed

    Nguyen, Scott V; McShan, William M

    2014-01-01

    Streptococcus pyogenes is a significant pathogen of humans, annually causing over 700,000,000 infections and 500,000 deaths. Virulence in S. pyogenes is closely linked to mobile genetic elements like phages and chromosomal islands (CI). S. pyogenes phage-like chromosomal islands (SpyCI) confer a complex mutator phenotype on their host. SpyCI integrate into the 5' end of DNA mismatch repair (MMR) gene mutL, which also disrupts downstream operon genes lmrP, ruvA, and tag. During early logarithmic growth, SpyCI excise from the bacterial chromosome and replicate as episomes, relieving the mutator phenotype. As growth slows and the cells enter stationary phase, SpyCI reintegrate into the chromosome, again silencing the MMR operon. This system creates a unique growth-dependent and reversible mutator phenotype. Additional CI using the identical attachment site in mutL have been identified in related species, including Streptococcus dysgalactiae subsp. equisimilis, Streptococcus anginosus, Streptococcus intermedius, Streptococcus parauberis, and Streptococcus canis. These CI have small genomes, which range from 13 to 20 kB, conserved integrase and DNA replication genes, and no identifiable genes encoding capsid proteins. SpyCI may employ a helper phage for packaging and dissemination in a fashion similar to the Staphylococcus aureus pathogenicity islands (SaPI). Outside of the core replication and integration genes, SpyCI and related CI show considerable diversity with the presence of many indels that may contribute to the host cell phenotype or fitness. SpyCI are a subset of a larger family of streptococcal CI who potentially regulate the expression of other host genes. The biological and phylogenetic analysis of streptococcal chromosomal islands provides important clues as to how these chromosomal islands help S. pyogenes and other streptococcal species persist in human populations in spite of antibiotic therapy and immune challenges.

  18. Chromosomal islands of Streptococcus pyogenes and related streptococci: molecular switches for survival and virulence

    PubMed Central

    Nguyen, Scott V.; McShan, William M.

    2014-01-01

    Streptococcus pyogenes is a significant pathogen of humans, annually causing over 700,000,000 infections and 500,000 deaths. Virulence in S. pyogenes is closely linked to mobile genetic elements like phages and chromosomal islands (CI). S. pyogenes phage-like chromosomal islands (SpyCI) confer a complex mutator phenotype on their host. SpyCI integrate into the 5′ end of DNA mismatch repair (MMR) gene mutL, which also disrupts downstream operon genes lmrP, ruvA, and tag. During early logarithmic growth, SpyCI excise from the bacterial chromosome and replicate as episomes, relieving the mutator phenotype. As growth slows and the cells enter stationary phase, SpyCI reintegrate into the chromosome, again silencing the MMR operon. This system creates a unique growth-dependent and reversible mutator phenotype. Additional CI using the identical attachment site in mutL have been identified in related species, including Streptococcus dysgalactiae subsp. equisimilis, Streptococcus anginosus, Streptococcus intermedius, Streptococcus parauberis, and Streptococcus canis. These CI have small genomes, which range from 13 to 20 kB, conserved integrase and DNA replication genes, and no identifiable genes encoding capsid proteins. SpyCI may employ a helper phage for packaging and dissemination in a fashion similar to the Staphylococcus aureus pathogenicity islands (SaPI). Outside of the core replication and integration genes, SpyCI and related CI show considerable diversity with the presence of many indels that may contribute to the host cell phenotype or fitness. SpyCI are a subset of a larger family of streptococcal CI who potentially regulate the expression of other host genes. The biological and phylogenetic analysis of streptococcal chromosomal islands provides important clues as to how these chromosomal islands help S. pyogenes and other streptococcal species persist in human populations in spite of antibiotic therapy and immune challenges. PMID:25161960

  19. The early interferon response of nasal-associated lymphoid tissue to Streptococcus pyogenes infection.

    PubMed

    Hyland, Kendra A; Brennan, Robert; Olmsted, Stephen B; Rojas, Eduardo; Murphy, Ellen; Wang, Beinan; Cleary, P Patrick

    2009-04-01

    Streptococcus pyogenes is a major causative agent of tonsillitis or pharyngitis in children. Streptococcus pyogenes can persist in tonsils, and one-third of children treated with antibiotics continue to shed streptococci and have recurrent infections. Mouse nasal-associated lymphoid tissue (NALT) is functionally analogous to human oropharyngeal lymphoid tissues, and serves as a model for characterization of the mucosal innate immune response to S. pyogenes. Wild-type S. pyogenes induces transcription of both type I and interferon-gamma (IFN-gamma)-responsive genes, proinflammatory genes and acute-phase response proteins 24 h after intranasal infection. Invasion of NALT and the induction of the interferon response were not dependent on expression of antiphagocytic M protein. Intranasal infection induces a substantial influx of neutrophils into NALT at 24 h, which declines by 48 h after infection. Infection of IFN-gamma(-/-) [IFN-gamma knock-out mouse (GKO)] C57BL/6 mice with wild-type S. pyogenes resulted in local dissemination of bacteria to draining lymph nodes (LN), but did not lead to systemic infection by 48 h after infection. Infected GKO mice had an increased influx of neutrophils into NALT compared with immunocompetent mice. Thus, IFN-gamma-induced responses are required to prevent local dissemination of streptococci to the draining LN.

  20. Rapid detection of Streptococcus pyogenes in pediatric patient specimens by DNA probe.

    PubMed Central

    Steed, L L; Korgenski, E K; Daly, J A

    1993-01-01

    A chemiluminescent DNA probe test (Group A Streptococcus Direct Test; Gen-Probe, Inc., San Diego, Calif.) for rapid, direct detection of cRNA of Streptococcus pyogenes in throat swabs was compared with conventional culture and identification techniques. Throat swabs from 277 patients suspected of having streptococcal pharyngitis were examined. By DNA probe alone, 10 specimens were positive, 51 were positive by both assays, and 8 were positive by culture alone. Thus, DNA probe sensitivity, specificity, and positive and negative predictive values were 86, 95, 84, and 96%, respectively. Including an indeterminate category, sensitivity, specificity, and positive and negative predictive values were 89, 96, 86, and 97%, respectively. After discrepancy testing, these values for the raw data improved to 90, 98, 93, and 97%, respectively. None of the 24 specimens that grew non-S. pyogenes beta-hemolytic streptococci in culture were positive by the DNA probe. Because mucoid S. pyogenes strains are more virulent than nonmucoid strains, 24 isolates were retrospectively tested with the DNA probe to ensure that both types would be detected equally well. Isolates were examined in pure cultures as well as mixed with representative normal oral flora. There was no statistical difference in detection of any of the four groups. Group A Streptococcus Direct Test is a rapid, sensitive, and specific test for S. pyogenes. PMID:8263185

  1. drs (Distantly Related sic) Gene Polymorphisms among emm12-Type Streptococcus pyogenes Isolates

    PubMed Central

    Brandt, Claudia M.; Haase, Gerhard; Spellerberg, Barbara; Holland, Regina; Lütticken, Rudolf

    2003-01-01

    Twenty-eight emm12-type Streptococcus pyogenes isolates from patients with invasive and noninvasive infections or from asymptomatic carriers were genetically typed. Sequencing of drs (distantly related sic [streptococcal inhibitor of complement]) genes identified two novel alleles and revealed a polymorphism for drs similar to that of sic. No association was observed between the five different drs alleles and the five restriction patterns of the vir regulon for the isolates studied. These data suggest that drs sequencing may be useful for further differentiation of S. pyogenes isolates with emm12 and identical vir regulon restriction patterns. PMID:12682191

  2. Complete Genome Sequence of Streptococcus pyogenes emm14 JS95, a Necrotizing Fasciitis Strain Isolated in Israel

    PubMed Central

    Chee, Jacqueline L. Y.; Ravins, Miriam; Hanski, Emanuel

    2017-01-01

    ABSTRACT Here, we report the complete genome sequence of the Streptococcus pyogenes emm14 strain JS95, isolated from a patient with necrotizing fasciitis. The streptococcal invasion locus (sil), the first quorum-sensing system characterized in S. pyogenes, was identified in this strain. PMID:28302774

  3. Complete Genome Sequence of Streptococcus pyogenes emm14 JS95, a Necrotizing Fasciitis Strain Isolated in Israel.

    PubMed

    Chee, Jacqueline L Y; Ravins, Miriam; Hanski, Emanuel; Chen, Swaine L

    2017-03-16

    Here, we report the complete genome sequence of the Streptococcus pyogenes emm14 strain JS95, isolated from a patient with necrotizing fasciitis. The streptococcal invasion locus (sil), the first quorum-sensing system characterized in S. pyogenes, was identified in this strain.

  4. Rapid detection of Streptococcus pyogenes in throat swab specimens by fluorescent in situ hybridization.

    PubMed

    Tajbakhsh, S; Gharibi, S; Zandi, K; Yaghobi, R; Asayesh, G

    2011-03-01

    Streptococcus pyogenes (S. pyogenes) is an important cause of pharyngitis. Rapid detection of this microorganism in throat specimens is essential to promptly start antibiotic therapy which could be lead to prevent complications and stop transmission of infection to other individuals. In the present study, fluorescent in situ hybridization (FISH) was compared with culture method for the detection of S. pyogenes in throat swab specimens. One hundred eleven patients with pharyngitis were included in this study. The throat swab specimens of these patients were investigated by both conventional culturing and FISH. Based on the results of this investigation, the sensitivity and specificity of FISH were 88.9% and 97.8%, respectively. Strikingly, in the specimen of one patient who had received antibiotic previous to clinical sampling, S. pyogenes was detected by means of FISH, whereas the culture method could not detect this bacterium. It seems that FISH is a suitable method for quick identification of S. pyogenes in throat swab specimens. When FISH is positive, culturing is not necessary. But because of the limited sensitivity of FISH for detection of S. pyogenes in throat swab specimens, culturing shoud be performed if FISH was negative.

  5. Six-Month Multicenter Study on Invasive Infections Due to Streptococcus pyogenes and Streptococcus dysgalactiae subsp. equisimilis in Argentina

    PubMed Central

    Lopardo, Horacio A.; Vidal, Patricia; Sparo, Monica; Jeric, Paola; Centron, Daniela; Facklam, Richard R.; Paganini, Hugo; Pagniez, N. Gaston; Lovgren, Marguerite; Beall, Bernard

    2005-01-01

    During a 6-month period, 95 invasive infections due to Streptococcus pyogenes and group C or group G Streptococcus dysgalactiae subsp. equisimilis were recorded from 40 centers of 16 cities in Argentina. We describe here epidemiologic data available for 55 and 19 patients, respectively, associated with invasive infections due to S. pyogenes and S. dysgalactiae subsp. equisimilis. The associated isolates and 58 additional pharyngeal isolates were genotyped and subjected to serologic and/or antibiotic susceptibility testing. Group A streptococcal emm type distribution and strain association with toxic shock appeared to differ somewhat from results found within the United States; however, serologic characterization and sof sequence typing suggested that emm types found in both countries are reflective of shared clonal types. PMID:15695683

  6. Increase in fluoroquinolone non-susceptibility among clinical Streptococcus pyogenes in Belgium during 2007-10.

    PubMed

    Van Heirstraeten, Liesbet; Leten, Gert; Lammens, Christine; Goossens, Herman; Malhotra-Kumar, Surbhi

    2012-11-01

    To study the temporal evolution of fluoroquinolone non-susceptibility among Streptococcus pyogenes during 2007-10 in Belgium. S. pyogenes (n = 4690) recovered from patients with tonsillopharyngitis or skin, wound or invasive infections were screened for fluoroquinolone non-susceptibility. A selection of fluoroquinolone-non-susceptible strains was investigated for resistance mechanisms: reserpine-sensitive efflux and mutations in topoisomerase genes parC and gyrA. Clonality was determined by emm typing. Fluoroquinolone non-susceptibility (ciprofloxacin MIC ≥2 mg/L) was identified in 535 (11.4%) of 4690 S. pyogenes recovered during 2007-10 in Belgium. The proportion of fluoroquinolone-non-susceptible S. pyogenes increased significantly from 4.3% (2008) to 10.9% (2009) to 21.6% (2010) and coincided with a significant increase in emm6 strains among fluoroquinolone-non-susceptible S. pyogenes. Ciprofloxacin MICs of 2-8 mg/L correlated with first-step ParC substitutions. Two high-level fluoroquinolone-resistant S. pyogenes strains (ciprofloxacin MICs 32 mg/L) showed second-step substitutions in GyrA (Ser-81→Phe or Tyr) in addition to first-step mutations in parC. Reserpine-sensitive efflux was not observed. We report an unprecedented increase in fluoroquinolone-non-susceptible S. pyogenes in Belgium, a country with high quinolone use, as well as emergence of two high-level fluoroquinolone-resistant S. pyogenes strains with second-step mutations in gyrA, warning us of the need for more prudent use of fluoroquinolones and for continued resistance surveillance.

  7. Effect of subinhibitory concentrations of fluoroquinolones on biofilm production by clinical isolates of Streptococcus pyogenes.

    PubMed

    Balaji, Kannan; Thenmozhi, Ramalingam; Pandian, Shunmugiah Karutha

    2013-05-01

    Subinhibitory concentrations (sub-MICs) of antibiotics, although not able to kill bacteria, but influence bacterial virulence significantly. Fluoroquinolones (FQs) which are used against other bacterial pathogens creates resistance in non-targeted Streptococcus pyogenes. This study was undertaken to characterize the effect of sub-MICs of FQs on S. pyogenes biofilm formation. Biofilm forming six M serotypes M56, st38, M89, M65, M100 and M74 of S. pyogenes clinical isolates were challenged against four FQs namely, ciprofloxacin, ofloxacin, levofloxacin and norfloxacin. The antibiofilm potential of these FQs was analysed at their subinhibitory concentrations (1/2 to 1/64 MIC) using biofilm assay, XTT reduction assay, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Among the four FQs tested, ofloxacin and levofloxacin at 1/2 MIC showed the maximum inhibition (92%) of biofilm formation against M56 and M74 serotypes. FQs effectively interfered in the microcolony formation of S. pyogenes isolates at 1/2 to 1/8 sub-MICs. Inhibition of biofilm formation was greatly reduced beyond 1/16 MICs and allowed biofilm formation. XTT reduction assay revealed the increase in metabolic activity of S. pyogenes biofilm against the decrease in FQs concentration. SEM and CLSM validated the potential of sub-MICs of FQs against the six S. pyogenes. Our results showed that the inhibitory effect all four FQs on S. pyogenes biofilm formation was concentration dependent. FQs at proper dosage can be effective against S. pyogenes and lower concentrations may allow the bacteria to form barriers against the antibiotic in the form of biofilm.

  8. Effect of subinhibitory concentrations of fluoroquinolones on biofilm production by clinical isolates of Streptococcus pyogenes

    PubMed Central

    Balaji, Kannan; Thenmozhi, Ramalingam; Pandian, Shunmugiah Karutha

    2013-01-01

    Background & objectives: Subinhibitory concentrations (sub-MICs) of antibiotics, although not able to kill bacteria, but influence bacterial virulence significantly. Fluoroquinolones (FQs) which are used against other bacterial pathogens creates resistance in non-targeted Streptococcus pyogenes. This study was undertaken to characterize the effect of sub-MICs of FQs on S. pyogenes biofilm formation. Methods: Biofilm forming six M serotypes M56, st38, M89, M65, M100 and M74 of S. pyogenes clinical isolates were challenged against four FQs namely, ciprofloxacin, ofloxacin, levofloxacin and norfloxacin. The antibiofilm potential of these FQs was analysed at their subinhibitory concentrations (1/2 to 1/64 MIC) using biofilm assay, XTT reduction assay, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Results: Among the four FQs tested, ofloxacin and levofloxacin at 1/2 MIC showed the maximum inhibition (92%) of biofilm formation against M56 and M74 serotypes. FQs effectively interfered in the microcolony formation of S. pyogenes isolates at 1/2 to 1/8 sub-MICs. Inhibition of biofilm formation was greatly reduced beyond 1/16 MICs and allowed biofilm formation. XTT reduction assay revealed the increase in metabolic activity of S. pyogenes biofilm against the decrease in FQs concentration. SEM and CLSM validated the potential of sub-MICs of FQs against the six S. pyogenes. Interpretation & conclusions: Our results showed that the inhibitory effect all four FQs on S. pyogenes biofilm formation was concentration dependent. FQs at proper dosage can be effective against S. pyogenes and lower concentrations may allow the bacteria to form barriers against the antibiotic in the form of biofilm. PMID:23760384

  9. Streptococcus pyogenes Infection in a Free-Living European Hedgehog (Erinaceus europaeus).

    PubMed

    Franklinos, Lydia H V; Efstratiou, Androulla; Macgregor, Shaheed K; John, Shinto K; Hopkins, Timothy; Cunningham, Andrew A; Lawson, Becki

    2015-12-01

    Streptococcus pyogenes, a common pathogen of humans, was isolated from the carcass of a free-living European hedgehog (Erinaceus europaeus) found in northern England in June 2014. The animal had abscessation of the deep right cervical lymph node, mesenteric lymph nodes and liver. The S. pyogenes strain isolated from the lesions, peritoneal and pleural cavities was characterised as emm 28, which can be associated with invasive disease in humans. This is the first known report of S. pyogenes in a hedgehog and in any free-living wild animal that has been confirmed by gene sequencing. As close associations between wild hedgehogs and people in England are common, we hypothesise that this case might have resulted from anthroponotic infection.

  10. A Novel Erythromycin Resistance Methylase Gene (ermTR) in Streptococcus pyogenes

    PubMed Central

    Seppälä, Helena; Skurnik, Mikael; Soini, Hanna; Roberts, Marilyn C.; Huovinen, Pentti

    1998-01-01

    Erythromycin resistance among streptococci is commonly due to target site modification by an rRNA-methylating enzyme, which results in coresistance to macrolide, lincosamide, and streptogramin B antibiotics (MLSB resistance). Genes belonging to the ermAM (ermB) gene class are the only erythromycin resistance methylase (erm) genes in Streptococcus pyogenes with MLSB resistance that have been sequenced so far. We identified a novel erm gene, designated ermTR, from an erythromycin-resistant clinical strain of S. pyogenes (strain A200) with an inducible type of MLSB resistance. The nucleotide sequence of ermTR is 82.5% identical to ermA, previously found, for example, in Staphylococcus aureus and coagulase-negative staphylococci. Our finding provides the first sequence of an erm gene other than ermAM that mediates MLSB resistance in S. pyogenes. PMID:9527769

  11. Genome Analysis of Streptococcus pyogenes Associated with Pharyngitis and Skin Infections

    PubMed Central

    Ibrahim, Joe; Eisen, Jonathan A.; Jospin, Guillaume; Coil, David A.; Khazen, Georges

    2016-01-01

    Streptococcus pyogenes is a very important human pathogen, commonly associated with skin or throat infections but can also cause life-threatening situations including sepsis, streptococcal toxic shock syndrome, and necrotizing fasciitis. Various studies involving typing and molecular characterization of S. pyogenes have been published to date; however next-generation sequencing (NGS) studies provide a comprehensive collection of an organism’s genetic variation. In this study, the genomes of nine S. pyogenes isolates associated with pharyngitis and skin infection were sequenced and studied for the presence of virulence genes, resistance elements, prophages, genomic recombination, and other genomic features. Additionally, a comparative phylogenetic analysis of the isolates with global clones highlighted their possible evolutionary lineage and their site of infection. The genomes were found to also house a multitude of features including gene regulation systems, virulence factors and antimicrobial resistance mechanisms. PMID:27977735

  12. The role of streptokinase as a virulence determinant of Streptococcus pyogenes--potential for therapeutic targeting.

    PubMed

    McArthur, Jason D; Cook, Simon M; Venturini, Carola; Walker, Mark J

    2012-03-01

    Streptococcus pyogenes is a major human pathogen responsible for numerous diseases ranging from uncomplicated skin and throat infections to severe, life threatening invasive disease such as necrotising fasciitis and streptococcal toxic shock syndrome. These severe invasive infections progress rapidly and produce high rates of morbidity and mortality despite the implementation of aggressive treatment plans. The activation of plasminogen and the acquisition of plasmin activity at the bacterial cell surface is critical for the invasive pathogenesis of this organism. To facilitate this process, S. pyogenes secrete streptokinase, a potent plasminogen activating protein. Here, we describe the role of streptokinase in invasive pathogenesis and discuss some potentially useful strategies for disruption of streptokinase mediated plasminogen activation which could be employed to treat severe invasive S. pyogenes infections.

  13. Genome Analysis of Streptococcus pyogenes Associated with Pharyngitis and Skin Infections.

    PubMed

    Ibrahim, Joe; Eisen, Jonathan A; Jospin, Guillaume; Coil, David A; Khazen, Georges; Tokajian, Sima

    2016-01-01

    Streptococcus pyogenes is a very important human pathogen, commonly associated with skin or throat infections but can also cause life-threatening situations including sepsis, streptococcal toxic shock syndrome, and necrotizing fasciitis. Various studies involving typing and molecular characterization of S. pyogenes have been published to date; however next-generation sequencing (NGS) studies provide a comprehensive collection of an organism's genetic variation. In this study, the genomes of nine S. pyogenes isolates associated with pharyngitis and skin infection were sequenced and studied for the presence of virulence genes, resistance elements, prophages, genomic recombination, and other genomic features. Additionally, a comparative phylogenetic analysis of the isolates with global clones highlighted their possible evolutionary lineage and their site of infection. The genomes were found to also house a multitude of features including gene regulation systems, virulence factors and antimicrobial resistance mechanisms.

  14. Flow cytometric assessment of susceptibilities of Streptococcus pyogenes to erythromycin and rokitamycin.

    PubMed

    Braga, Pier Carlo; Bovio, Cinzia; Culici, Maria; Dal Sasso, Monica

    2003-01-01

    The effects of erythromycin (a 14-membered ring macrolide) and rokitamycin (a 16-membered ring macrolide) on the viability of the Streptococcus pyogenes M phenotype were studied by means of flow cytometry and fluorescence microscopy by using a combination of two fluorochromes (syto 9 and propidium iodide) that stains live bacteria green and dead bacteria red. In order to apply the flow cytometry, a bacterial sonication procedure was expressly set up to separate single cells from the long, intralaced S. pyogenes chains of up to 30 to 40 cells that have previously prevented the application of flow cytometry to this type of bacteria. The association of flow cytometry using an appropriate sonication procedure, together with a combination of fluorescent probes, offered the possibility of very quickly investigating the different microbiological effects of rokitamycin at 2 microg/ml, which was active on the S. pyogenes M phenotype, and of erythromycin at doses of up to 32 microg/ml, which was not.

  15. An examination of the differential sensitivity to ketolide antibiotics in ermB strains of Streptococcus pyogenes and Streptococcus pneumoniae.

    PubMed

    Champney, W Scott; Mentens, Nicole; Zurawick, Kimberly

    2004-10-01

    Several reports in the literature have described a differential sensitivity to ketolide antibiotics in ermB strains of Streptococcus pyogenes and Streptococcus pneumoniae resistant to erythromycin. Strains of S. pyogenes and S. pneumoniae carrying different erm gene alleles were examined for their susceptibility to the ketolide antibiotics cethromycin (ABT-773) and telithromycin. The effect of the antibiotics on cell growth and viability was assessed as were effects on protein synthesis and 50S ribosomal subunit formation. The susceptibility of wild-type strains of both organisms was compared with effects in strains containing the ermA and ermB methyltransferase genes. A wild-type antibiotic-susceptible strain of S. pyogenes was comparable to an ermA strain of the organism in its ketolide sensitivity, with IC(50) values for 50% inhibition of protein synthesis and 50S ribosomal subunit formation of 10 ng/mL for cethromycin and 16 ng/mL for telithromycin. An S. pneumoniae strain with the ermB gene and an S. pyogenes strain with the ermA gene were also similar in their sensitivity to ketolide inhibition. IC(50) values for inhibition of translation and subunit formation in S. pneumoniae ( ermB) were 30 ng/mL and 55 ng/mL and for the ermA strain of S. pyogenes they were 15 ng/mL and 35 ng/mL respectively. By contrast, an S. pyogenes ermB strain was significantly more resistant to both ketolides, with IC(50) values for inhibition of 50S synthesis of 215 and 380 ng/mL for the two ketolides. Experiments were conducted to examine ribosome synthesis and translational activity in the two ermB strains at intervals during growth in the presence of each antibiotic. Cell viability and 50S subunit formation were dramatically reduced in the S. pneumoniae strain during continued growth with either drug. By contrast, the ketolides had little effect on the S. pyogenes strain growing with the antibiotics. The results indicate that ketolides have a reduced inhibitory effect on

  16. Genetic resistance elements carrying mef subclasses other than mef(A) in Streptococcus pyogenes.

    PubMed

    Del Grosso, Maria; Camilli, Romina; Barbabella, Giada; Blackman Northwood, John; Farrell, David J; Pantosti, Annalisa

    2011-07-01

    In Streptococcus pyogenes, efflux-mediated erythromycin resistance is associated with the mef gene, represented mostly by mef(A), although a small portion of strains carry different mef subclasses. We characterized the composite genetic elements, including mef subclasses other than mef(A), associated with other resistance genes in S. pyogenes isolates. Determination of the genetic elements was performed by PCR mapping. The strains carrying mosaic mef(A/E), in which the 5' region was identical to mef(A) and the 3' region was identical to mef(E), also carried tet(O). The two genes were found enclosed in an element similar to S. pyogenes prophage Φm46.1, designated the Φm46.1-like element. In S. pyogenes strains carrying mef(E) and tet(M), mef(E) was included in a typical mega element, and in some strains, it was physically associated with tet(M) in the composite element Tn2009. S. pyogenes strains carrying mef(I) also carried catQ; the two genes were linked in a fragment representing a portion of the 5216IQ complex of Streptococcus pneumoniae, designated the defective IQ element. In the only isolate carrying a novel mef gene, this was associated with catQ and tet(M) in a genetic element similar to the 5216IQ complex of S. pneumoniae (5216IQ-like complex), suggesting that the novel mef is in fact a variant of mef(I). This study demonstrates that the composite elements containing mef are shared between S. pyogenes and S. pneumoniae and suggests that it is important to distinguish the mef subclass on the basis of the genetic element containing it.

  17. Clinical and Microbiologic Characteristics of Invasive Streptococcus pyogenes Infections in North and South India

    PubMed Central

    Haggar, Axana; Nerlich, Andreas; Kumar, Rajesh; Abraham, Vinod J.; Brahmadathan, Kootallur N.; Ray, Pallab; Dhanda, Vanita; Joshua, John Melbin Jose; Mehra, Narinder; Bergmann, Rene; Chhatwal, G. Singh

    2012-01-01

    The lack of epidemiologic data on invasive Streptococcus pyogenes infections in many developing countries is concerning, as S. pyogenes infections are commonly endemic in these areas. Here we present the results of the first prospective surveillance study of invasive Streptococcus pyogenes infections in India. Fifty-four patients with invasive S. pyogenes infections were prospectively enrolled at two study sites, one in the north and one in the south of India. Sterile-site isolates were collected, and clinical information was documented using a standardized questionnaire. Available acute-phase sera were tested for their ability to inhibit superantigens produced by the patient's own isolate using a cell-based neutralizing assay. The most common clinical presentations were bacteremia without focus (30%), pneumonia (28%), and cellulitis (17%). Only two cases of streptococcal toxic shock syndrome and no cases of necrotizing fasciitis were identified. Characterization of the isolates revealed great heterogeneity, with 32 different emm subtypes and 29 different superantigen gene profiles being represented among the 49 sterile-site isolates. Analyses of acute-phase sera showed that only 20% of the cases in the north cohort had superantigen-neutralizing activity in their sera, whereas 50% of the cases from the south site had neutralizing activity. The results demonstrate that there are important differences in both clinical presentation and strain characteristics between invasive S. pyogenes infections in India and invasive S. pyogenes infections in Western countries. The findings underscore the importance of epidemiologic studies on streptococcal infections in India and have direct implications for current vaccine developments. PMID:22357508

  18. Purification, crystallization and preliminary crystallographic analysis of Streptococcus pyogenes laminin-binding protein Lbp

    SciTech Connect

    Linke, Christian; Caradoc-Davies, Tom T.; Proft, Thomas; Baker, Edward N.

    2008-02-01

    The S. pyogenes laminin-binding protein Lbp, which is essential for adhesion to human laminin, has been expressed, purified and crystallized. The laminin-binding protein Lbp (Spy2007) from Streptococcus pyogenes (a group A streptococcus) mediates adhesion to the human basal lamina glycoprotein laminin. Accordingly, Lbp is essential in in vitro models of cell adhesion and invasion. However, the molecular and structural basis of laminin binding by bacteria remains unknown. Therefore, the lbp gene has been cloned for recombinant expression in Escherichia coli. Lbp has been purified and crystallized from 30%(w/v) PEG 1500 by the sitting-drop vapour-diffusion method. The crystals belonged to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 42.62, b = 92.16, c = 70.61 Å, β = 106.27°, and diffracted to 2.5 Å resolution.

  19. [Epidemiology and clinical features of Streptococcus pyogenes bacteremia in Cartagena (Murcia, Spain)].

    PubMed

    Jimeno-Almazán, Amaya; Viqueira-Gonzalez, Montserrat; Alcalde, María Del Mar; Alcaraz-Vidal, Begoña; Vera-Méndez, Francisco

    2013-01-01

    A gradual increase in severe cases due to Streptococcus pyogenes or Streptococcus beta-hemolytic group A (SGA), has been detected in the last few decades. Retrospective study of bacteremia due to S.pyogenes detected between January 2009 and January 2013 in Cartagena. The annual incidence for severe bacteremia has been estimated. Thirteen cases of SGA bacteremia were recorded. The incidence increased from 0.37 in 2009 to 2.5 cases/100,000 inhabitants in 2012. The predominant focus was skin and soft tissue infections (53%). Early mortality was 20%. Severe streptococcal disease is rare, but affects individuals with good functional status, and is associated with a high mortality. Copyright © 2012 Elsevier España, S.L. All rights reserved.

  20. PURIFICATION AND PROPERTIES OF N-ACETYL-d-GLUCOSAMINE KINASE FROM STREPTOCOCCUS PYOGENES

    PubMed Central

    Zeleznick, L. D.; Hankin, H.; Boltralik, J. J.; Heymann, H.; Barkulis, S. S.

    1964-01-01

    Zeleznick, L. D. (CIBA Pharmaceutical Co., Summit, N.J.), H. Hankin, J. J. Boltralik, H. Heymann, and S. S. Barkulis. Purification and properties of N-acetyl-d-glucosamine kinase from Streptococcus pyogenes. J. Bacteriol. 88:1288–1295. 1964.—A kinase from Streptococcus pyogenes which catalyzes adenosine triphosphate-dependent phosphorylation of d-glucose and N-acetyl-d-glucosamine has been purified 1,500-fold. The ratio of the enzymatic activity on both substrates remained constant throughout the fractionation. Similarity in heat stability, p-hydroxymercuribenzoate inhibition, protection by either carbohydrate, and lack of repression of enzymatic activity when the bacteria were grown exclusively on one of the two substrates supports the hypothesis that kinase activity is associated with one enzyme. PMID:14234783

  1. [Streptococcus pyogenes or group A streptococcal infections in child: French national reference center data].

    PubMed

    Bidet, P; Plainvert, C; Doit, C; Mariani-Kurkdjian, P; Bonacorsi, S; Lepoutre, A; Bouvet, A; Poyart, C; Bingen, E

    2010-02-01

    Since the 1980s, infections due to Streptococcus pyogenes or group A streptococci (GAS) were marked by the increase in invasive infections and the emergence of clones which were resistant to macrolides. Those challenges led the French national reference center for streptococci to enhance the epidemiological survey and the characterization of GAS strains, in collaboration with the National Institute for Public Health Surveillance. Active surveillance is of major importance for implementation of therapeutic and prophylactic guidelines and for evaluation of future streptococcal vaccines.

  2. Growth phase-dependent effect of clindamycin on production of exoproteins by Streptococcus pyogenes.

    PubMed

    Sawai, Jun; Hasegawa, Tadao; Kamimura, Takuya; Okamoto, Akira; Ohmori, Daisuke; Nosaka, Nobuyuki; Yamada, Keiko; Torii, Keizo; Ohta, Michio

    2007-02-01

    The administration of high-dose clindamycin plus benzylpenicillin has been recommended for the treatment of streptococcal toxic shock-like syndrome caused by Streptococcus pyogenes, and clindamycin has been found to be more effective than beta-lactams in retrospective analyses of human cases. Although therapeutic doses of clindamycin have also been shown to be effective against experimental infections and clindamycin has great efficacy against the production of bacterial exoproteins, we recently reported that the level of production of some exoproteins was unchanged or even increased by a subinhibitory dose of clindamycin when it is added upon the initiation of bacterial culture and the treated cultures were analyzed by two-dimensional gel electrophoresis. In this study we further examined the effect of clindamycin on the production of exoproteins by adding it to Streptococcus pyogenes cultures during various growth phases. We found that the levels of production of some proteins, NAD+ glycohydrolase, streptolysin O, and streptococcal inhibitor of complement, were increased when clindamycin was added at early-log-phase growth, which was the result that was seen when clindamycin was added at the beginning of culture. However, clindamycin inhibited the production of most types of proteins when it was administered to Streptococcus pyogenes cultures at mid-log-phase growth. In csrS- or mga-knockout bacterial strains, the increase in exoproteins seen in parental strains was considerably inhibited. Our study indicates that the in vitro effect of clindamycin on the production of exoproteins greatly depends on the growth phase of bacteria and some regulatory factors of Streptococcus pyogenes that are involved in this phenomenon.

  3. Getting under the skin: the immunopathogenesis of Streptococcus pyogenes deep tissue infections.

    PubMed

    Johansson, Linda; Thulin, Pontus; Low, Donald E; Norrby-Teglund, Anna

    2010-07-01

    Streptococcus pyogenes can cause a variety of diseases in immunocompetent individuals, from pharyngotonsillitis to life-threatening invasive diseases, such as streptococcal toxic shock syndrome, and rapidly progressing deep-tissue infections, such as necrotizing fasciitis. Necrotizing fasciitis is often seen in combination with streptococcal toxic shock syndrome, which further increases morbidity and mortality. We review here the host-pathogen interactions in the tissue milieu and discuss the use of intravenous immunoglobulin as potential adjunctive therapy in these life-threatening infections.

  4. Recurrent Streptococcus pyogenes genital infection in a woman: test and treat the partner!

    PubMed

    Verkaeren, Emilienne; Epelboin, Loïc; Epelboin, Sylvie; Boddaert, Nathalie; Brossier, Florence; Caumes, Eric

    2014-12-01

    Group A Streptococcus (GAS) is a well-known cause of vulvovaginitis in prepubescent girls, but it is rarely described in adult women. We describe the case of a 64-year-old woman who presented with endometritis revealed by GAS bacteraemia, followed by recurrent vulvovaginitis due to a wild-type strain of GAS. She relapsed twice despite amoxicillin treatment. Her husband was found to be an asymptomatic carrier after GAS was identified in nasal and rectal swabs. She was cured after eradication of carriage in both herself and her husband with amoxicillin and rifampin. When recurrent Streptococcus pyogenes genital infections occur, test and treat the partner.

  5. Kinetics of cytokine profile in response to Mycobacterium bovis BCG and Streptococcus pyogenes activated cells.

    PubMed

    Verma, Vivek; Kumar, Parveen; Dhanda, Rakesh Singh; Yadav, Manisha

    2016-06-01

    The infection of epithelial cells is a necessary step for Mycobacterium bovis BCG dissemination, but the mechanism of mycobacterial epithelial interactions is not completely understood. Similarly, Streptococcus pyogenes is a strictly human pathogen that favorably colonizes the skin and the pharynx. Effective cytokine secretion is essential in order to fabricate a suitable inflammatory response against an infection. In this data article, the cytokine profile in BCG and S. pyogenes activated THP-1 cell line in media after the acute phase of infection by ELISA is described. The interleukin-8 level was increased in response to both BCG and S. pyogenes, but was quite prominent after 24 h and further increased upto 72 h post infection. On the other hand, an increase in IL-6 response to S. pyogenes was observed while there was no response to BCG even after 48 h of infection. A low level of TNF-α was detected upon BCG and S. pyogenes infection.

  6. Induction of Cyclooxygenase 2 by Streptococcus pyogenes Is Mediated by Cytolysins.

    PubMed

    Blaschke, Ulrike; Beineke, Andreas; Klemens, Johanna; Medina, Eva; Goldmann, Oliver

    2017-08-17

    Prostaglandin E2 (PGE2), an arachidonic acid metabolite regulating a broad range of physiological activities, is an important modulator of the severity of infection caused by Streptococcus pyogenes. Here, we investigated the role of streptococcal cytolysin S (SLS) and streptococcal cytolysin O (SLO) in the induction of cyclooxygenase-2 (COX-2), the rate-limiting enzyme in the synthesis of prostaglandins, in in vitro cultured macrophages and during in vivo infection. Macrophages were infected with S. pyogenes wild type or with the isogenic mutant strains deficient in SLS (ΔSLS), SLO (ΔSLO), or both (ΔSLS/ΔSLO), and the expression of COX-2 was determined at the transcriptional and the protein level. The results indicated that S. pyogenes induced expression of COX-2 and concomitant synthesis of PGE2 in macrophages mediated by the synergistic activity of both SLS and SLO, and involved calcium and the PKC/JNK signaling pathway. These results were validated using recombinant cytolysins. In a murine skin infection model, COX-2-positive cells were found more abundant at the site of S. pyogenes wild-type infection than at the site of infection with ΔSLS/ΔSLO mutant strain. These findings suggest that inhibitory targeting of SLS and SLO could ameliorate the adverse effects of high levels of prostaglandins during S. pyogenes infection. © 2017 S. Karger AG, Basel.

  7. Streptococcus pyogenes CAMP factor attenuates phagocytic activity of RAW 264.7 cells.

    PubMed

    Kurosawa, Mie; Oda, Masataka; Domon, Hisanori; Saitoh, Issei; Hayasaki, Haruaki; Terao, Yutaka

    2016-02-01

    Streptococcus pyogenes produces molecules that inhibit the function of human immune system, thus allowing the pathogen to grow and spread in tissues. It is known that S. pyogenes CAMP factor increases erythrocytosis induced by Staphylococcus aureus β-hemolysin. However, the effects of CAMP factor for immune cells are unclear. In this study, we investigated the effects of CAMP factor to macrophages. Western blotting analysis demonstrated that all examined strains expressed CAMP factor protein. In the presence of calcium or magnesium ion, CAMP factor was significantly released in the supernatant. In addition, both culture supernatant from S. pyogenes strain SSI-9 and recombinant CAMP factor dose-dependently induced vacuolation in RAW 264.7 cells, but the culture supernatant from Δcfa isogenic mutant strain did not. CAMP factor formed oligomers in RAW 264.7 cells in a time-dependent manner. CAMP factor suppressed cell proliferation via G2 phase cell cycle arrest without inducing cell death. Furthermore, CAMP factor reduced the uptake of S. pyogenes and phagocytic activity indicator by RAW 264.7 cells. These results suggest that CAMP factor works as a macrophage dysfunction factor. Therefore, we conclude that CAMP factor allows S. pyogenes to escape the host immune system, and contribute to the spread of streptococcal infection. Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  8. mef(A) is the predominant macrolide resistance determinant in Streptococcus pneumoniae and Streptococcus pyogenes in Germany.

    PubMed

    Bley, Christine; van der Linden, Mark; Reinert, Ralf René

    2011-05-01

    In this study, macrolide-resistant Streptococcus pneumoniae and Streptococcus pyogenes isolates from Germany were carefully characterised by susceptibility testing, phenotyping, polymerase chain reaction (PCR) and sequencing of macrolides resistance genes, and multilocus sequence typing (MLST). Of 2045 S. pneumoniae and 352 S. pyogenes isolates, 437 (21.4%) and 29 (8.2%), respectively, were found to be macrolide-resistant. Amongst the S. pneumoniae isolates, the most prevalent resistance marker was mef(A) (57.7%) followed by erm(B) (27.0%) and mef(E) (11.2%). Of note, the dual resistance mechanism mef(E)+erm(B) was found in a relatively high proportion (4.1%) of pneumococcal isolates. Amongst the S. pyogenes isolates, 31.0% carried mef(A), 34.5% erm(B) and 13.8% erm(A). Dissemination of a single clone [mef(A)-positive England(14)-9] has significantly contributed to the emergence of macrolide resistance amongst pneumococci in Germany.

  9. Kinetic characterization of arginine deiminase and carbamate kinase from Streptococcus pyogenes M49.

    PubMed

    Hering, Silvio; Sieg, Antje; Kreikemeyer, Bernd; Fiedler, Tomas

    2013-09-01

    Streptococcus pyogenes (group A Streptococcus, GAS) is an important human pathogen causing mild superficial infections of skin and mucous membranes, but also life-threatening systemic diseases. S. pyogenes and other prokaryotic organisms use the arginine deiminase system (ADS) for survival in acidic environments. In this study, the arginine deiminase (AD), and carbamate kinase (CK) from S. pyogenes M49 strain 591 were heterologously expressed in Escherichia coli DH5α, purified, and kinetically characterized. AD and CK from S. pyogenes M49 share high amino acid sequence similarity with the respective enzymes from Lactococcus lactis subsp. lactis IL1403 (45.6% and 53.5% identical amino acids) and Enterococcus faecalis V583 (66.8% and 66.8% identical amino acids). We found that the arginine deiminase of S. pyogenes is not allosterically regulated by the intermediates and products of the arginine degradation (e.g., ATP, citrulline, carbamoyl phosphate). The Km and Vmax values for arginine were 1.13±0.12mM (mean±SD) and 1.51±0.07μmol/min/mg protein. The carbamate kinase is inhibited by ATP but unaffected by arginine and citrulline. The Km and Vmax values for ADP were 0.72±0.08mM and 1.10±0.10μmol/min/mg protein and the Km for carbamoyl phosphate was 0.65±0.07mM. The optimum pH and temperature for both enzymes were 6.5 and 37°C, respectively.

  10. ICESpy009, a Conjugative Genetic Element Carrying mef(E) in Streptococcus pyogenes.

    PubMed

    Del Grosso, Maria; Camilli, Romina; Rizzi, Ermanno; Pietrelli, Alessandro; De Bellis, Gianluca; Pantosti, Annalisa

    2016-07-01

    Efflux-mediated macrolide resistance due to mef(E) and mel, carried by the mega element, is common in Streptococcus pneumoniae, for which it was originally characterized, but it is rare in Streptococcus pyogenes In S. pyogenes, mega was previously found to be enclosed in Tn2009, a composite genetic element of the Tn916 family containing tet(M) and conferring erythromycin and tetracycline resistance. In this study, S. pyogenes isolates containing mef(E), apparently not associated with other resistance determinants, were examined to characterize the genetic context of mega. By whole-genome sequencing of one isolate, MB56Spyo009, we identified a novel composite integrative and conjugative element (ICE) carrying mega, designated ICESpy009, belonging to the ICESa2603 family. ICESpy009 was 55 kb long, contained 61 putative open reading frames (ORFs), and was found to be integrated into hylA, a novel integration site for the ICESa2603 family. The modular organization of the ICE was similar to that of members of the ICESa2603 family carried by different streptococcal species. In addition, a novel cluster of accessory resistance genes was found inside a region that encloses mega. PCR mapping targeting ICESpy009 revealed the presence of a similar ICE in five other isolates under study. While in three isolates the integration site was the same as that of ICESpy009, in two isolates the ICE was integrated into rplL, the typical integration site of the ICESa2603 family. ICESpy009 was able to transfer macrolide resistance by conjugation to both S. pyogenes and S. pneumoniae, showing the first evidence of the transferability of mega from S. pyogenes.

  11. ICESpy009, a Conjugative Genetic Element Carrying mef(E) in Streptococcus pyogenes

    PubMed Central

    Camilli, Romina; Rizzi, Ermanno; Pietrelli, Alessandro; De Bellis, Gianluca; Pantosti, Annalisa

    2016-01-01

    Efflux-mediated macrolide resistance due to mef(E) and mel, carried by the mega element, is common in Streptococcus pneumoniae, for which it was originally characterized, but it is rare in Streptococcus pyogenes. In S. pyogenes, mega was previously found to be enclosed in Tn2009, a composite genetic element of the Tn916 family containing tet(M) and conferring erythromycin and tetracycline resistance. In this study, S. pyogenes isolates containing mef(E), apparently not associated with other resistance determinants, were examined to characterize the genetic context of mega. By whole-genome sequencing of one isolate, MB56Spyo009, we identified a novel composite integrative and conjugative element (ICE) carrying mega, designated ICESpy009, belonging to the ICESa2603 family. ICESpy009 was 55 kb long, contained 61 putative open reading frames (ORFs), and was found to be integrated into hylA, a novel integration site for the ICESa2603 family. The modular organization of the ICE was similar to that of members of the ICESa2603 family carried by different streptococcal species. In addition, a novel cluster of accessory resistance genes was found inside a region that encloses mega. PCR mapping targeting ICESpy009 revealed the presence of a similar ICE in five other isolates under study. While in three isolates the integration site was the same as that of ICESpy009, in two isolates the ICE was integrated into rplL, the typical integration site of the ICESa2603 family. ICESpy009 was able to transfer macrolide resistance by conjugation to both S. pyogenes and S. pneumoniae, showing the first evidence of the transferability of mega from S. pyogenes. PMID:27067338

  12. Macrolide-Resistant Streptococcus pneumoniae and Streptococcus pyogenes in the Pediatric Population in Germany during 2000-2001

    PubMed Central

    Reinert, Ralf René; Lütticken, Rudolf; Bryskier, André; Al-Lahham, Adnan

    2003-01-01

    In a nationwide study in Germany covering 13 clinical microbiology laboratories, a total of 307 Streptococcus pyogenes (mainly pharyngitis) and 333 Streptococcus pneumoniae (respiratory tract infections) strains were collected from outpatients less than 16 years of age. The MICs of penicillin G, amoxicillin, cefotaxime, erythromycin A, clindamycin, levofloxacin, and telithromycin were determined by the microdilution method. In S. pyogenes isolates, resistance rates were as follows: penicillin, 0%; erythromycin A, 13.7%; and levofloxacin, 0%. Telithromycin showed good activity against S. pyogenes isolates (MIC90 = 0.25 μg/ml; MIC range, 0.016 to 16 μg/ml). Three strains were found to be telithromycin-resistant (MIC ≥ 4 μg/ml). Erythromycin-resistant strains were characterized for the underlying resistance genotype, with 40.5% having the efflux type mef(A), 38.1% having the erm(A), and 9.5% having the erm(B) genotypes. emm typing of macrolide-resistant S. pyogenes isolates showed emm types 4 (45.2%), 77 (26.2%), and 12 (11.9%) to be predominant. In S. pneumoniae, resistance rates were as follows: penicillin intermediate, 7.5%; penicillin resistant, 0%; erythromycin A, 17.4%; and levofloxacin, 0%. Telithromycin was highly active against pneumococcal isolates (MIC90 ≤ 0.016 μg/ml; range, 0.016 to 0.5 μg/ml). The overall resistance profile of streptococcal respiratory tract isolates is still favorable, but macrolide resistance is of growing concern in Germany. PMID:12543648

  13. Molecular Analysis of an Outbreak of Lethal Postpartum Sepsis Caused by Streptococcus pyogenes

    PubMed Central

    Turner, Claire E.; Dryden, Matthew; Holden, Matthew T. G.; Davies, Frances J.; Lawrenson, Richard A.; Farzaneh, Leili; Bentley, Stephen D.; Efstratiou, Androulla

    2013-01-01

    Sepsis is now the leading direct cause of maternal death in the United Kingdom, and Streptococcus pyogenes is the leading pathogen. We combined conventional and genomic analyses to define the duration and scale of a lethal outbreak. Two postpartum deaths caused by S. pyogenes occurred within 24 h; one was characterized by bacteremia and shock and the other by hemorrhagic pneumonia. The women gave birth within minutes of each other in the same maternity unit 2 days earlier. Seven additional infections in health care and household contacts were subsequently detected and treated. All cluster-associated S. pyogenes isolates were genotype emm1 and were initially indistinguishable from other United Kingdom emm1 isolates. Sequencing of the virulence gene sic revealed that all outbreak isolates had the same unique sic type. Genome sequencing confirmed that the cluster was caused by a unique S. pyogenes clone. Transmission between patients occurred on a single day and was associated with casual contact only. A single isolate from one patient demonstrated a sequence change in sic consistent with longer infection duration. Transmission to health care workers was traced to single clinical contacts with index cases. The last case was detected 18 days after the first case. Following enhanced surveillance, the outbreak isolate was not detected again. Mutations in bacterial regulatory genes played no detectable role in this outbreak, illustrating the intrinsic ability of emm1 S. pyogenes to spread while retaining virulence. This fast-moving outbreak highlights the potential of S. pyogenes to cause a range of diseases in the puerperium with rapid transmission, underlining the importance of immediate recognition and response by clinical infection and occupational health teams. PMID:23616448

  14. Resistance to multiple fluoroquinolones in a clinical isolate of Streptococcus pyogenes: identification of gyrA and parC and specification of point mutations associated with resistance.

    PubMed

    Yan, S S; Fox, M L; Holland, S M; Stock, F; Gill, V J; Fedorko, D P

    2000-11-01

    A strain of Streptococcus pyogenes resistant to multiple fluoroquinolones was isolated from the blood of an immunocompromised patient. Resistance to fluoroquinolones in S. pyogenes has not been previously studied. Compared to 10 sensitive strains of S. pyogenes, the fluoroquinolone-resistant clinical isolate of S. pyogenes presented point mutations in gyrA, predicting that serine-81 was changed to phenylalanine and that methionine-99 was changed to leucine, and in parC, predicting that serine-79 was changed to tyrosine. The mechanism of fluoroquinolone resistance in this isolate of S. pyogenes appears to be analogous to previously reported mechanisms for Streptococcus pneumoniae.

  15. Resistance to Multiple Fluoroquinolones in a Clinical Isolate of Streptococcus pyogenes: Identification of gyrA and parC and Specification of Point Mutations Associated with Resistance

    PubMed Central

    Yan, S. Steve; Fox, Melissa L.; Holland, Steven M.; Stock, Frida; Gill, Vee J.; Fedorko, Daniel P.

    2000-01-01

    A strain of Streptococcus pyogenes resistant to multiple fluoroquinolones was isolated from the blood of an immunocompromised patient. Resistance to fluoroquinolones in S. pyogenes has not been previously studied. Compared to 10 sensitive strains of S. pyogenes, the fluoroquinolone-resistant clinical isolate of S. pyogenes presented point mutations in gyrA, predicting that serine-81 was changed to phenylalanine and that methionine-99 was changed to leucine, and in parC, predicting that serine-79 was changed to tyrosine. The mechanism of fluoroquinolone resistance in this isolate of S. pyogenes appears to be analogous to previously reported mechanisms for Streptococcus pneumoniae. PMID:11036052

  16. Characterization of levofloxacin non-susceptible clinical Streptococcus pyogenes isolated in the central part of Italy.

    PubMed

    Petrelli, D; Di Luca, M C; Prenna, M; Bernaschi, P; Repetto, A; Vitali, L A

    2014-02-01

    We investigated the prevalence, genetics, and clonality of fluoroquinolone non-susceptible isolates of Streptococcus pyogenes in the central part of Italy. S. pyogenes strains (n = 197) were isolated during 2012 from patients with tonsillopharyngitis, skin, wound or invasive infections and screened for fluoroquinolone non-susceptibility (resistance to norfloxacin and levofloxacin minimum inhibitory concentration (MIC) = 2 mg/L) following EUCAST guidelines. First-step topoisomerase parC and gyrA substitutions were investigated using sequencing analysis. Clonality was determined by pulsed field gel electrophoresis (PFGE; SmaI digestion) and by emm typing. The fluoroquinolone non-susceptible phenotype was identified in 18 isolates (9.1 %) and correlated with mutations in parC, but not in gyrA, the most frequent leading to substitution of the serine at position 79 with an alanine. Most of the fluoroquinolone non-susceptible isolates belonged to the emm-type 6, even if other emm-types were also represented (emm75, emm89, and emm2). A significant level of association was measured between PFGE and both emm type and substitutions in parC. The prevalence of fluoroquinolone non-susceptible Streptococcus pyogenes isolates in Italy is of concern and, although the well-known emm type 6 is dominant, other types are appearing and spreading.

  17. NrdI essentiality for class Ib ribonucleotide reduction in Streptococcus pyogenes.

    PubMed

    Roca, Ignasi; Torrents, Eduard; Sahlin, Margareta; Gibert, Isidre; Sjöberg, Britt-Marie

    2008-07-01

    The Streptococcus pyogenes genome harbors two clusters of class Ib ribonucleotide reductase genes, nrdHEF and nrdF*I*E*, and a second stand-alone nrdI gene, designated nrdI2. We show that both clusters are expressed simultaneously as two independent operons. The NrdEF enzyme is functionally active in vitro, while the NrdE*F* enzyme is not. The NrdF* protein lacks three of the six highly conserved iron-liganding side chains and cannot form a dinuclear iron site or a tyrosyl radical. In vivo, on the other hand, both operons are functional in heterologous complementation in Escherichia coli. The nrdF*I*E* operon requires the presence of the nrdI* gene, and the nrdHEF operon gained activity upon cotranscription of the heterologous nrdI gene from Streptococcus pneumoniae, while neither nrdI* nor nrdI2 from S. pyogenes rendered it active. Our results highlight the essential role of the flavodoxin NrdI protein in vivo, and we suggest that it is needed to reduce met-NrdF, thereby enabling the spontaneous reformation of the tyrosyl radical. The NrdI* flavodoxin may play a more direct role in ribonucleotide reduction by the NrdF*I*E* system. We discuss the possibility that the nrdF*I*E* operon has been horizontally transferred to S. pyogenes from Mycoplasma spp.

  18. Novel Bacteriophage Lysin with Broad Lytic Activity Protects against Mixed Infection by Streptococcus pyogenes and Methicillin-Resistant Staphylococcus aureus

    PubMed Central

    Gilmer, Daniel B.; Schmitz, Jonathan E.; Euler, Chad W.

    2013-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) and Streptococcus pyogenes (group A streptococcus [GrAS]) cause serious and sometimes fatal human diseases. They are among the many Gram-positive pathogens for which resistance to leading antibiotics has emerged. As a result, alternative therapies need to be developed to combat these pathogens. We have identified a novel bacteriophage lysin (PlySs2), derived from a Streptococcus suis phage, with broad lytic activity against MRSA, vancomycin-intermediate S. aureus (VISA), Streptococcus suis, Listeria, Staphylococcus simulans, Staphylococcus epidermidis, Streptococcus equi, Streptococcus agalactiae (group B streptococcus [GBS]), S. pyogenes, Streptococcus sanguinis, group G streptococci (GGS), group E streptococci (GES), and Streptococcus pneumoniae. PlySs2 has an N-terminal cysteine-histidine aminopeptidase (CHAP) catalytic domain and a C-terminal SH3b binding domain. It is stable at 50°C for 30 min, 37°C for >24 h, 4°C for 15 days, and −80°C for >7 months; it maintained full activity after 10 freeze-thaw cycles. PlySs2 at 128 μg/ml in vitro reduced MRSA and S. pyogenes growth by 5 logs and 3 logs within 1 h, respectively, and exhibited a MIC of 16 μg/ml for MRSA. A single, 2-mg dose of PlySs2 protected 92% (22/24) of the mice in a bacteremia model of mixed MRSA and S. pyogenes infection. Serially increasing exposure of MRSA and S. pyogenes to PlySs2 or mupirocin resulted in no observed resistance to PlySs2 and resistance to mupirocin. To date, no other lysin has shown such notable broad lytic activity, stability, and efficacy against multiple, leading, human bacterial pathogens; as such, PlySs2 has all the characteristics to be an effective therapeutic. PMID:23571534

  19. RNA sequencing uncovers antisense RNAs and novel small RNAs in Streptococcus pyogenes

    PubMed Central

    Le Rhun, Anaïs; Beer, Yan Yan; Reimegård, Johan; Chylinski, Krzysztof; Charpentier, Emmanuelle

    2016-01-01

    ABSTRACT Streptococcus pyogenes is a human pathogen responsible for a wide spectrum of diseases ranging from mild to life-threatening infections. During the infectious process, the temporal and spatial expression of pathogenicity factors is tightly controlled by a complex network of protein and RNA regulators acting in response to various environmental signals. Here, we focus on the class of small RNA regulators (sRNAs) and present the first complete analysis of sRNA sequencing data in S. pyogenes. In the SF370 clinical isolate (M1 serotype), we identified 197 and 428 putative regulatory RNAs by visual inspection and bioinformatics screening of the sequencing data, respectively. Only 35 from the 197 candidates identified by visual screening were assigned a predicted function (T-boxes, ribosomal protein leaders, characterized riboswitches or sRNAs), indicating how little is known about sRNA regulation in S. pyogenes. By comparing our list of predicted sRNAs with previous S. pyogenes sRNA screens using bioinformatics or microarrays, 92 novel sRNAs were revealed, including antisense RNAs that are for the first time shown to be expressed in this pathogen. We experimentally validated the expression of 30 novel sRNAs and antisense RNAs. We show that the expression profile of 9 sRNAs including 2 predicted regulatory elements is affected by the endoribonucleases RNase III and/or RNase Y, highlighting the critical role of these enzymes in sRNA regulation. PMID:26580233

  20. Synergy and Mode of Action of Ceftazidime plus Quercetin or Luteolin on Streptococcus pyogenes

    PubMed Central

    Siriwong, Supatcharee; Thumanu, Kanjana; Hengpratom, Tanaporn; Eumkeb, Griangsak

    2015-01-01

    Streptococcus pyogenes causes streptococcal toxic shock syndrome. The recommended therapy has been often failure through the interfering of beta-lactamase-producing bacteria (BLPB). The present study was to investigate antibacterial activity, synergy, and modes of action of luteolin and quercetin using alone and plus ceftazidime against S. pyogenes. The MICs of ceftazidime, luteolin, and quercetin against all S. pyogenes were 0.50, 128, and 128 µg mL−1, respectively. A synergistic effect was exhibited on luteolin and quercetin plus ceftazidime against these strains at fractional inhibitory concentration indices 0.37 and 0.27, respectively, and was confirmed by the viable count. These combinations increased cytoplasmic membrane (CM) permeability, caused irregular cell shape, peptidoglycan, and CM damage, and decreased nucleic acid but increased proteins in bacterial cells. Enzyme assay demonstrated that these flavonoids had an inhibitory activity against β-lactamase. In summary, this study provides evidence that the inhibitory mode of action of luteolin and quercetin may be mediated via three mechanisms: (1) inhibiting of peptidoglycan synthesis, (2) increasing CM permeability, and (3) decreasing nucleic acid but increasing the protein contents of bacterial cells. So, luteolin and quercetin propose the high potential to develop adjunct to ceftazidime for the treatment of coexistence of the BLPB and S. pyogenes infections. PMID:26576195

  1. Antibacterial Activity of Rhodomyrtus tomentosa (Aiton) Hassk. Leaf Extract against Clinical Isolates of Streptococcus pyogenes.

    PubMed

    Limsuwan, Surasak; Kayser, Oliver; Voravuthikunchai, Supayang Piyawan

    2012-01-01

    Ethanol extract of Rhodomyrtus tomentosa (Aiton) Hassk. leaf was evaluated for antibacterial activity against 47 clinical isolates of Streptococcus pyogenes. The extract exhibited good anti-S. pyogenes activity against all the tested isolates with similar minimum inhibitory concentration (MIC, 3.91-62.5 μg mL(-1)) and minimum bactericidal concentration (MBC, 3.91-62.5 μg mL(-1)) ranges. No surviving cells were detected at 16 h after treatment with 8 × MIC of the extract. The extract-treated cells demonstrated no lysis and cytoplasmic leakage through the bacterial membrane. Electron micrographs further revealed that the extract did not cause any dramatic changes on the treated cells. Rhodomyrtone, an isolated compound, exhibited good anti-S. pyogenes activity (14 isolates), expressed very low MIC (0.39-1.56 μg mL(-1)) and MBC (0.39-1.56 μg mL(-1)) values. Rhodomyrtus tomentosa leaf extract and rhodomyrtone displayed promising antibacterial activity against clinical isolates of S. pyogenes.

  2. Inhibition of Growth and Gene Expression by PNA-peptide Conjugates in Streptococcus pyogenes

    PubMed Central

    Patenge, Nadja; Pappesch, Roberto; Krawack, Franziska; Walda, Claudia; Mraheil, Mobarak Abu; Jacob, Anette; Hain, Torsten; Kreikemeyer, Bernd

    2013-01-01

    While Streptococcus pyogenes is consistently susceptible toward penicillin, therapeutic failure of penicillin treatment has been reported repeatedly and a considerable number of patients exhibit allergic reactions to this substance. At the same time, streptococcal resistance to alternative antibiotics, e.g., macrolides, has increased. Taken together, these facts demand the development of novel therapeutic strategies. In this study, S. pyogenes growth was inhibited by application of peptide-conjugated antisense-peptide nucleic acids (PNAs) specific for the essential gyrase A gene (gyrA). Thereby, HIV-1 Tat peptide-coupled PNAs were more efficient inhibitors of streptococcal growth as compared with (KFF)3K-coupled PNAs. Peptide-anti-gyrA PNAs decreased the abundance of gyrA transcripts in S. pyogenes. Growth inhibition by antisense interference was enhanced by combination of peptide-coupled PNAs with protein-level inhibitors. Antimicrobial synergy could be detected with levofloxacin and novobiocin, targeting the gyrase enzyme, and with spectinomycin, impeding ribosomal function. The prospective application of carrier peptide-coupled antisense PNAs in S. pyogenes covers the use as an antimicrobial agent and the employment as a knock-down strategy for the investigation of virulence factor function. PMID:24193033

  3. Antibacterial Activity of Rhodomyrtus tomentosa (Aiton) Hassk. Leaf Extract against Clinical Isolates of Streptococcus pyogenes

    PubMed Central

    Limsuwan, Surasak; Kayser, Oliver; Voravuthikunchai, Supayang Piyawan

    2012-01-01

    Ethanol extract of Rhodomyrtus tomentosa (Aiton) Hassk. leaf was evaluated for antibacterial activity against 47 clinical isolates of Streptococcus pyogenes. The extract exhibited good anti-S. pyogenes activity against all the tested isolates with similar minimum inhibitory concentration (MIC, 3.91–62.5 μg mL−1) and minimum bactericidal concentration (MBC, 3.91–62.5 μg mL−1) ranges. No surviving cells were detected at 16 h after treatment with 8 × MIC of the extract. The extract-treated cells demonstrated no lysis and cytoplasmic leakage through the bacterial membrane. Electron micrographs further revealed that the extract did not cause any dramatic changes on the treated cells. Rhodomyrtone, an isolated compound, exhibited good anti-S. pyogenes activity (14 isolates), expressed very low MIC (0.39–1.56 μg mL−1) and MBC (0.39-1.56 μg mL−1) values. Rhodomyrtus tomentosa leaf extract and rhodomyrtone displayed promising antibacterial activity against clinical isolates of S. pyogenes. PMID:22973404

  4. Mucosal vaccine made from live, recombinant Lactococcus lactis protects mice against pharyngeal infection with Streptococcus pyogenes.

    PubMed

    Mannam, Praveen; Jones, Kevin F; Geller, Bruce L

    2004-06-01

    A novel vaccine (LL-CRR) made from live, nonpathogenic Lactococcus lactis that expresses the conserved C-repeat region (CRR) of M protein from Streptococcus pyogenes serotype 6 was tested in mice. Nasally vaccinated mice produced CRR-specific salivary immunoglobulin A (IgA) and serum IgG. Subcutaneously vaccinated mice produced CRR-specific serum IgG but not salivary IgA. A combined regimen produced responses similar to the salivary IgA of nasally vaccinated mice and serum IgG of subcutaneously vaccinated mice. Mice vaccinated nasally or with the combined regimen were significantly protected against pharyngeal infection following a nasal challenge with S. pyogenes M serotype 14. Mice vaccinated subcutaneously were not protected against pharyngeal infection. Mice in all three LL-CRR vaccination groups were significantly protected against the lethal effects of S. pyogenes. Only 1 of 77 challenged mice that were vaccinated with LL-CRR died, whereas 60 of 118 challenged mice that were vaccinated with a control strain or phosphate-buffered saline died. In conclusion, mucosal vaccination with LL-CRR produced CRR-specific salivary IgA and serum IgG, prevented pharyngeal infection with S. pyogenes, and promoted survival.

  5. Pili mediate specific adhesion of Streptococcus pyogenes to human tonsil and skin.

    PubMed

    Abbot, Emily L; Smith, Wendy D; Siou, Gerard P S; Chiriboga, Carlos; Smith, Rebecca J; Wilson, Janet A; Hirst, Barry H; Kehoe, Michael A

    2007-07-01

    Very little is known about the biological functions of pili that have recently been found to be expressed by important Gram-positive pathogens such as Corynebacterium diphtheriae, Streptococcus agalacticae, S. pneumoniae and S. pyogenes. Using various ex vivo tissue and cellular models, here we show that pili mediate adhesion of serotype M1 S. pyogenes strain SF370 to both human tonsil epithelium and primary human keratinocytes, which represent the two main sites of infection by this human-specific pathogen. Mutants lacking minor pilus subunits retained the ability to express cell-surface pili, but these were functionally defective. In contrast to above, pili were not required for S. pyogenes adhesion to either immortalized HEp-2 or A549 cells, highlighting an important limitation of these extensively used adhesion/invasion models. Adhering bacteria were internalized very effectively by both HEp-2 and A549 cells, but not by tonsil epithelium or primary keratinocytes. While pili acted as the primary adhesin, the surface M1 protein clearly enhanced adhesion to tonsil, but surprisingly, had the opposite effect on adhesion to keratinocytes. These studies provide clear evidence that S. pyogenes pili display an adhesive specificity for clinically relevant human tissues and are likely to play a critical role in the initial stages of infection.

  6. Streptococcus pyogenes cluster in a care home in England April to June 2010.

    PubMed

    Milne, L M; Lamagni, T; Efstratiou, A; Foley, C; Gilman, J; Lilley, M; Guha, S; Head, F; Han, T

    2011-11-24

    Two fatal cases of Streptococcus pyogenes emm st22.6 bacteraemia occurred in a care home in England during April and June 2010, initiating a cluster investigation. The first case had left the home 13 days before the second case took up residence. We sought further cases and carriers. We swabbed throat and chronic skin lesions from residents and staff and examined these specimens for the presence of S. pyogenes. 61 specimens were taken from 18 of 19 residents and 39 of 39 staff. All results from swabbing were culture negative. We observed infection control practices and the environment at the care home for deficiencies. Issues were identified relating to the correct use of personal protective equipment, hand hygiene, clinical waste and laundry. Infection control practices were improved and training given. Infection control practices and the environment at a care home should be examined as part of the investigation of a S. pyogenes cluster. Screening for carriage of S. pyogenes should be done before antibiotic chemoprophylaxis is issued to care home residents and staff.

  7. Transduction of the Streptococcus pyogenes bacteriophage Φm46.1, carrying resistance genes mef(A) and tet(O), to other Streptococcus species.

    PubMed

    Giovanetti, Eleonora; Brenciani, Andrea; Morroni, Gianluca; Tiberi, Erika; Pasquaroli, Sonia; Mingoia, Marina; Varaldo, Pietro E

    2014-01-01

    Φm46.1 - Streptococcus pyogenes bacteriophage carrying mef(A) and tet(O), respectively, encoding resistance to macrolides (M phenotype) and tetracycline - is widespread in S. pyogenes but has not been reported outside this species. Φm46.1 is transferable in vitro among S. pyogenes isolates, but no information is available about its transferability to other Streptococcus species. We thus investigated Φm46.1 for its ability to be transduced in vitro to recipients of different Streptococcus species. Transductants were obtained from recipients of Streptococcus agalactiae, Streptococcus gordonii, and Streptococcus suis. Retransfer was always achieved, and from S. suis to S. pyogenes occurred at a much greater frequency than in the opposite direction. In transductants Φm46.1 retained its functional properties, such as inducibility with mitomycin C, presence both as a prophage and as a free circular form, and transferability. The transductants shared the same Φm46.1 chromosomal integration site as the donor, at the 3' end of a conserved RNA uracil methyltransferase (rum) gene, which is an integration hotspot for a variety of genetic elements. No transfer occurred to recipients of Streptococcus pneumoniae, Streptococcus oralis, and Streptococcus salivarius, even though rum-like genes were also detected in the sequenced genomes of these species. A largely overlapping 18-bp critical sequence, where the site-specific recombination process presumably takes place, was identified in the rum genes of all recipients, including those of the species yielding no transductants. Growth assays to evaluate the fitness cost of Φm46.1 acquisition disclosed a negligible impact on S. pyogenes, S. agalactiae, and S. gordonii transductants and a noticeable fitness advantage in S. suis. The S. suis transductant also displayed marked overexpression of the autolysin-encoding gene atl.

  8. Antibodies against a Surface Protein of Streptococcus pyogenes Promote a Pathological Inflammatory Response

    PubMed Central

    Kahn, Fredrik; Mörgelin, Matthias; Shannon, Oonagh; Norrby-Teglund, Anna; Herwald, Heiko; Olin, Anders I.; Björck, Lars

    2008-01-01

    Streptococcal toxic shock syndrome (STSS) caused by Streptococcus pyogenes is a clinical condition with a high mortality rate despite modern intensive care. A key feature of STSS is excessive plasma leakage leading to hypovolemic hypotension, disturbed microcirculation and multiorgan failure. Previous work has identified a virulence mechanism in STSS where M1 protein of S. pyogenes forms complexes with fibrinogen that activate neutrophils to release heparin-binding protein (HBP), an inducer of vascular leakage. Here, we report a marked inter-individual difference in the response to M1 protein–induced HBP release, a difference found to be related to IgG antibodies directed against the central region of the M1 protein. To elicit massive HBP release, such antibodies need to be part of the M1 protein–fibrinogen complexes. The data add a novel aspect to bacterial pathogenesis where antibodies contribute to the severity of disease by promoting a pathologic inflammatory response. PMID:18787689

  9. Establishment of a superficial skin infection model in mice by using Staphylococcus aureus and Streptococcus pyogenes.

    PubMed

    Kugelberg, Elisabeth; Norström, Tobias; Petersen, Thomas K; Duvold, Tore; Andersson, Dan I; Hughes, Diarmaid

    2005-08-01

    A new animal model for the purpose of studying superficial infections is presented. In this model an infection is established by disruption of the skin barrier by partial removal of the epidermal layer by tape stripping and subsequent application of the pathogens Staphylococcus aureus and Streptococcus pyogenes. The infection and the infection route are purely topical, in contrast to those used in previously described animal models in mice, such as the skin suture-wound model, where the infection is introduced into the deeper layers of the skin. Thus, the present model is considered more biologically relevant for the study of superficial skin infections in mice and humans. Established topical antibiotic treatments are shown to be effective. The procedures involved in the model are simple, a feature that increases throughput and reproducibility. This new model should be applicable to the evaluation of novel antimicrobial treatments of superficial infections caused by S. aureus and S. pyogenes.

  10. Antibiotic Susceptibility of Streptococcus Pyogenes Isolated from Respiratory Tract Infections in Dakar, Senegal

    PubMed Central

    Camara, Makhtar; Dieng, Assane; Boye, Cheikh Saad Bouh

    2013-01-01

    Group A Streptococcus (GAS) is one of the major causes of respiratory tract infections. The objectives of this study were to identify isolates of S. pyogenes obtained from respiratory tract infections, and to assess their susceptibility to several antibiotics. A total of 40 strains were isolated and their susceptibility to 17 antibiotics was tested using a standard disk diffusion method. The minimum inhibitory concentrations (MICs) were determined using the E-test. All isolates were sensitive to β-lactam antibiotics including penicillin, amoxicillin, and cephalosporins. Macrolides remain active with the exception of spiramycin, which showed reduced susceptibility. Out of the 40 isolates, 100% of the isolates were resistant to tetracycline. Interestingly, isolates were sensitive to chloramphenicol, teicoplanin, vancomycine, and levofloxacin, providing potential alternative choices of treatment against infections with S. pyogenes. PMID:24826076

  11. Typing of Streptococcus pyogenes strains isolated from throat infections in the region of Aachen, Germany.

    PubMed

    Brandt, C M; Spellerberg, B; Honscha, M; Truong, N D; Hoevener, B; Lütticken, R

    2001-01-01

    Changes in the epidemiology of Streptococcus pyogenes infections may be associated with the introduction and reappearance of individual serotypes within a population. Typing of 216 consecutive isolates of S. pyogenes from patients with pharyngitis in the region of Aachen, Germany, was performed by sequencing the emm gene, slide-agglutination of the T-antigen and determining the serum opacity reaction (SOR). All 216 isolates were unequivocally emm-typable. emm1 was most common (18.5%), foLlowed by emm12 (15.7%), emm3 (14.4%) and emm28 (13.9%). Only four isolates contained newly validated emm types: emm89 or emm94 were harbored by two isolates each. In one isolate, the sequence type s104 was found. Despite an anticipated selective pressure, the prevalence of emm1 among isolates from throat infections in northwestern Germany remains high, but does not reflect the predominance of emm1 among invasive isolates in Germany.

  12. Survival of Streptococcus suis, Streptococcus dysgalactiae and Trueperella pyogenes in dry-cured Iberian pork shoulders and loins.

    PubMed

    Cardoso-Toset, F; Luque, I; Morales-Partera, A; Galán-Relaño, A; Barrero-Domínguez, B; Hernández, M; Gómez-Laguna, J

    2017-02-01

    Dry-cured hams, shoulders and loins of Iberian pigs are highly appreciated in national and international markets. Salting, additive addition and dehydration are the main strategies to produce these ready-to-eat products. Although the dry curing process is known to reduce the load of well-known food borne pathogens, studies evaluating the viability of other microorganisms in contaminated pork have not been performed. In this work, the efficacy of the dry curing process to eliminate three swine pathogens associated with pork carcass condemnation, Streptococcus suis, Streptococcus dysgalactiae and Trueperella pyogenes, was evaluated. Results of this study highlight that the dry curing process is a suitable method to obtain safe ready-to-eat products free of these microorganisms. Although salting of dry-cured shoulders had a moderate bactericidal effect, results of this study suggest that drying and ripening were the most important stages to obtain dry-cured products free of these microorganisms.

  13. Antibacterial Activity of the Contact and Complement Systems Is Blocked by SIC, a Protein Secreted by Streptococcus pyogenes*

    PubMed Central

    Frick, Inga-Maria; Shannon, Oonagh; Åkesson, Per; Mörgelin, Matthias; Collin, Mattias; Schmidtchen, Artur; Björck, Lars

    2011-01-01

    Recent studies have shown that activation of complement and contact systems results in the generation of antibacterial peptides. Streptococcus pyogenes, a major bacterial pathogen in humans, exists in >100 different serotypes due to sequence variation in the surface-associated M protein. Cases of invasive and life-threatening S. pyogenes infections are commonly associated with isolates of the M1 serotype, and in contrast to the large majority of M serotypes, M1 isolates all secrete the SIC protein. Here, we show that SIC interferes with the activation of the contact system and blocks the activity of antibacterial peptides generated through complement and contact activation. This effect promotes the growth of S. pyogenes in human plasma, and in a mouse model of S. pyogenes sepsis, SIC enhances bacterial dissemination, results which help explain the high frequency of severe S. pyogenes infections caused by isolates of the M1 serotype. PMID:21068386

  14. Predominant role of msr(D) over mef(A) in macrolide resistance in Streptococcus pyogenes.

    PubMed

    Zhang, Yan; Tatsuno, Ichiro; Okada, Ryo; Hata, Nanako; Matsumoto, Masakado; Isaka, Masanori; Isobe, Ken-ichi; Hasegawa, Tadao

    2016-01-01

    In Japan, the number of patients with streptococcal toxic shock syndrome is reported to be increasing. mef(A) gene-positive macrolide-resistant emm1 strains are thought to possibly contribute to the rise in the frequency of STSS. Although analyses of macrolide-resistant mechanisms, including mef(A) resistance, have been performed mainly in Streptococcus pneumoniae, the role of this gene in Streptococcus pyogenes has not been completely investigated. Therefore, to the best of our knowledge, we established the first mef(A)-knockout strain using an emm1-type S. pyogenes strain, and tested its susceptibility to erythromycin, clarithromycin and azithromycin. We found that the antimicrobial susceptibilities were almost identical to those of the parental strain. Hence, we established a knockout strain for another gene, msr(D), that is located immediately downstream of mef(A). The macrolide resistances of the resulting strain significantly decreased, and were further altered when both mef(A) and msr(D) were knocked out. The introduction of the msr(D) gene into a macrolide-sensitive strain conferred more resistance than the introduction of the mef(A) gene. The erythromycin susceptibilities of knockout strains were further dissected using two additional emm4- and emm75-type S. pyogenes strains. We found almost identical results for both strains except for the mef(A) knockout emm4 type, whose susceptibility was altered, although the change was less than that for the msr(D) knockout. These results suggest that both mef(A) and msr(D) are involved in macrolide resistance in S. pyogenes, and that the msr(D) gene plays a more predominant role in macrolide resistance than mef(A).

  15. Local Th17/IgA immunity correlate with protection against intranasal infection with Streptococcus pyogenes.

    PubMed

    Mortensen, Rasmus; Christensen, Dennis; Hansen, Lasse Bøllehuus; Christensen, Jan Pravsgaard; Andersen, Peter; Dietrich, Jes

    2017-01-01

    Streptococcus pyogenes (group A streptococcus, GAS) is responsible for a wide array of infections. Respiratory transmission via droplets is the most common mode of transmission but it may also infect the host via other routes such as lesions in the skin. To advance the development of a future vaccine against GAS, it is therefore important to investigate how protective immunity is related to the route of vaccine administration. To explore this, we examined whether a parenterally administered anti-GAS vaccine could protect against an intranasal GAS infection or if this would require locally primed immunity. We foundd that a parenteral CAF01 adjuvanted GAS vaccine offered no protection against intranasal infection despite inducing strong systemic Th1/Th17/IgG immunity that efficiently protected against an intraperitoneal GAS infection. However, the same vaccine administered via the intranasal route was able to induce protection against repeated intranasal GAS infections in a murine challenge model. The lack of intranasal protection induced by the parenteral vaccine correlated with a reduced mucosal recall response at the site of infection. Taken together, our results demonstrate that locally primed immunity is important for the defense against intranasal infection with Streptococcus pyogenes.

  16. An Approach for Identification of Novel Drug Targets in Streptococcus pyogenes SF370 Through Pathway Analysis.

    PubMed

    Singh, Satendra; Singh, Dev Bukhsh; Singh, Anamika; Gautam, Budhayash; Ram, Gurudayal; Dwivedi, Seema; Ramteke, Pramod W

    2016-12-01

    Streptococcus pyogenes is one of the most important pathogens as it is involved in various infections affecting upper respiratory tract and skin. Due to the emergence of multidrug resistance and cross-resistance, S. Pyogenes is becoming more pathogenic and dangerous. In the present study, an in silico comparative analysis of total 65 metabolic pathways of the host (Homo sapiens) and the pathogen was performed. Initially, 486 paralogous enzymes were identified so that they can be removed from possible drug target list. The 105 enzymes of the biochemical pathways of S. pyogenes from the KEGG metabolic pathway database were compared with the proteins from the Homo sapiens by performing a BLASTP search against the non-redundant database restricted to the Homo sapiens subset. Out of these, 83 enzymes were identified as non-human homologous while 30 enzymes of inadequate amino acid length were removed for further processing. Essential enzymes were finally mined from remaining 53 enzymes. Finally, 28 essential enzymes were identified in S. pyogenes SF370 (serotype M1). In subcellular localization study, 18 enzymes were predicted with cytoplasmic localization and ten enzymes with the membrane localization. These ten enzymes with putative membrane localization should be of particular interest. Acyl-carrier-protein S-malonyltransferase, DNA polymerase III subunit beta and dihydropteroate synthase are novel drug targets and thus can be used to design potential inhibitors against S. pyogenes infection. 3D structure of dihydropteroate synthase was modeled and validated that can be used for virtual screening and interaction study of potential inhibitors with the target enzyme.

  17. Murine Vaginal Colonization Model for Investigating Asymptomatic Mucosal Carriage of Streptococcus pyogenes

    PubMed Central

    Watson, Michael E.; Nielsen, Hailyn V.; Hultgren, Scott J.

    2013-01-01

    While many virulence factors promoting Streptococcus pyogenes invasive disease have been described, specific streptococcal factors and host properties influencing asymptomatic mucosal carriage remain uncertain. To address the need for a refined model of prolonged S. pyogenes asymptomatic mucosal colonization, we have adapted a preestrogenized murine vaginal colonization model for S. pyogenes. In this model, derivatives of strains HSC5, SF370, JRS4, NZ131, and MEW123 established a reproducible, asymptomatic colonization of the vaginal mucosa over a period of typically 3 to 4 weeks' duration at a relatively high colonization efficiency. Prior treatment with estradiol prolonged streptococcal colonization and was associated with reduced inflammation in the colonized vaginal epithelium as well as a decreased leukocyte presence in vaginal fluid compared to the levels of inflammation and leukocyte presence in non-estradiol-treated control mice. The utility of our model for investigating S. pyogenes factors contributing to mucosal carriage was verified, as a mutant with a mutation in the transcriptional regulator catabolite control protein A (CcpA) demonstrated significant impairment in vaginal colonization. An assessment of in vivo transcriptional activity in the CcpA− strain for several known CcpA-regulated genes identified significantly elevated transcription of lactate oxidase (lctO) correlating with excessive generation of hydrogen peroxide to self-lethal levels. Deletion of lctO did not impair colonization, but deletion of lctO in a CcpA− strain prolonged carriage, exceeding even that of the wild-type strain. Thus, while LctO is not essential for vaginal colonization, its dysregulation is deleterious, highlighting the critical role of CcpA in promoting mucosal colonization. The vaginal colonization model should prove effective for future analyses of S. pyogenes mucosal colonization. PMID:23460515

  18. The structure of pyogenecin immunity protein, a novel bacteriocin-like immunity protein from streptococcus pyogenes.

    SciTech Connect

    Chang, C.; Coggill, P.; Bateman, A.; Finn, R.; Cymborowski, M.; Otwinowski, Z.; Minor, W.; Volkart, L.; Joachimiak, A.; Wellcome Trust Sanger Inst.; Univ. of Virginia; UT Southwestern Medical Center

    2009-12-17

    Many Gram-positive lactic acid bacteria (LAB) produce anti-bacterial peptides and small proteins called bacteriocins, which enable them to compete against other bacteria in the environment. These peptides fall structurally into three different classes, I, II, III, with class IIa being pediocin-like single entities and class IIb being two-peptide bacteriocins. Self-protective cognate immunity proteins are usually co-transcribed with these toxins. Several examples of cognates for IIa have already been solved structurally. Streptococcus pyogenes, closely related to LAB, is one of the most common human pathogens, so knowledge of how it competes against other LAB species is likely to prove invaluable. We have solved the crystal structure of the gene-product of locus Spy-2152 from S. pyogenes, (PDB: 2fu2), and found it to comprise an anti-parallel four-helix bundle that is structurally similar to other bacteriocin immunity proteins. Sequence analyses indicate this protein to be a possible immunity protein protective against class IIa or IIb bacteriocins. However, given that S. pyogenes appears to lack any IIa pediocin-like proteins but does possess class IIb bacteriocins, we suggest this protein confers immunity to IIb-like peptides. Combined structural, genomic and proteomic analyses have allowed the identification and in silico characterization of a new putative immunity protein from S. pyogenes, possibly the first structure of an immunity protein protective against potential class IIb two-peptide bacteriocins. We have named the two pairs of putative bacteriocins found in S. pyogenes pyogenecin 1, 2, 3 and 4.

  19. [Emergence of macrolide resistant Streptococcus pyogenes strains in pediatric patients in France].

    PubMed

    Mariani-Kurkdjian, P; Doit, C; Deforche, D; Brahimi, N; Francois, M; Van den Abbeele, T; Bingen, E

    2004-10-01

    A total of 206 recent throat isolates of Streptococcus pyogenes collected between 2002 and 2004 from children were tested for their susceptibility to penicillin, amoxycillin, erythromycin, clarythromycin and clindamycin. The erythromycin resistant isolates were further studied for their genetic mechanism of resistance by means of PCR. In all, 14.5% of the strains were erythromycin resistant and 13.5 and 1% expressed the constitutive MLS(B) and M resistance phenotypes and harbored the ermB and mef A genes respectively.

  20. Cutaneous bacterial infections caused by Staphylococcus aureus and Streptococcus pyogenes in infants and children.

    PubMed

    Larru, Beatriz; Gerber, Jeffrey S

    2014-04-01

    Acute bacterial skin and skin structure infections (SSSIs) are among the most common bacterial infections in children. The medical burden of SSSIs, particularly abscesses, has increased nationwide since the emergence of community-acquired methicillin-resistant Staphylococcus aureus. SSSIs represent a wide spectrum of disease severity. Prompt recognition, timely institution of appropriate therapy, and judicious antimicrobial use optimize patient outcomes. For abscesses, incision and drainage are paramount and might avoid the need for antibiotic treatment in uncomplicated cases. If indicated, empiric antimicrobial therapy should target Streptococcus pyogenes for nonpurulent SSSIs, such as uncomplicated cellulitis, and S aureus for purulent SSSIs such as abscesses. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Asymptomatic throat carriage rate and antimicrobial resistance pattern of Streptococcus pyogenes in Nepalese school children.

    PubMed

    Dumre, S P; Sapkota, K; Adhikari, N; Acharya, D; Karki, M; Bista, S; Basanyat, S R; Joshi, S K

    2009-01-01

    Streptococcus pyogenes or Group A streptococcus (GAS) causes several suppurative and non suppurative infections. In addition to pharyngitis and skin infections, GAS are also the causative agent of post-streptococcal infection syndromes such as acute rheumatic fever (ARF) and post-streptococcal glumerulonephritis (PSG). GAS frequently colonises in the throat of an asymptomatic person. Pharyngeal carriage rates of GAS among healthy school children vary with geographical location and seasons. We carried out this preliminary study to determine the throat carriage rate and antimicrobial resistance trend of Streptococcus pyogenes or Group A streptococcus (GAS) among the Nepalese school children. Four schools situated at different locations of Kathmandu valley were included in the study. Throat swabs from 350 students of age group 5-15 years were collected, immediately transported to the laboratory and were processed for S. pyogenes following standard microbiological procedures. Antimicrobial susceptibility testing of the isolates was performed by Kirby Bauer disc diffusion method following CLSI guidelines. S. pyogenes was isolated from 10.9% (38/350) of the screened children. The GAS colonisation rate was statistically insignificant (P>0.05) with sex and age sub-groups, although the rate was slightly higher among girls and age sub-group 9-12 years. No significant difference in carrier rate was observed among different schools (P>0.05). All isolates were susceptible to azithromycin. No resistance was detected for penicillin and its derivative antibiotic ampicillin. Highest resistance rate was observed for cotrimoxazole (71.0%) followed by chloramphenicol (7.8%), ciprofloxacin (5.2%) and erythromycin (5.2%). Antibiotic resistant GAS isolated from asymptomatic Nepalese school children is a public health concern. When screened and appropriately treated with antibiotics, carriers can be prevented from spreading of streptococcal infections in the school environment and the

  2. Streptococcus pyogenes degrades extracellular matrix in chondrocytes via MMP-13

    SciTech Connect

    Sakurai, Atsuo; Okahashi, Nobuo; Maruyama, Fumito; Ooshima, Takashi; Hamada, Shigeyuki; Nakagawa, Ichiro

    2008-08-29

    Group A streptococcus (GAS) causes a wide range of human diseases, including bacterial arthritis. The pathogenesis of arthritis is characterized by synovial proliferation and the destruction of cartilage and subchondral bone in joints. We report here that GAS strain JRS4 invaded a chondrogenic cell line ATDC5 and induced the degradation of the extracellular matrix (ECM), whereas an isogenic mutant of JRS4 lacking a fibronectin-binding protein, SAM1, failed to invade the chondrocytes or degrade the ECM. Reverse transcription-PCR and Western blot analysis revealed that the expression of matrix metalloproteinase (MMP)-13 was strongly elevated during the infection with GAS. A reporter assay revealed that the activation of the AP-1 transcription factor and the phosphorylation of c-Jun terminal kinase participated in MMP-13 expression. These results suggest that MMP-13 plays an important role in the destruction of infected joints during the development of septic arthritis.

  3. Endothelial cells are intrinsically defective in xenophagy of Streptococcus pyogenes

    PubMed Central

    Lu, Shiou-Ling; Kawabata, Tsuyoshi; Cheng, Yi-Lin; Omori, Hiroko; Hamasaki, Maho; Kusaba, Tatsuya; Iwamoto, Ryo; Lin, Yee-Shin

    2017-01-01

    Group A Streptococcus (GAS) is deleterious pathogenic bacteria whose interaction with blood vessels leads to life-threatening bacteremia. Although xenophagy, a special form of autophagy, eliminates invading GAS in epithelial cells, we found that GAS could survive and multiply in endothelial cells. Endothelial cells were competent in starvation-induced autophagy, but failed to form double-membrane structures surrounding GAS, an essential step in xenophagy. This deficiency stemmed from reduced recruitment of ubiquitin and several core autophagy proteins in endothelial cells, as demonstrated by the fact that it could be rescued by exogenous coating of GAS with ubiquitin. The defect was associated with reduced NO-mediated ubiquitin signaling. Therefore, we propose that the lack of efficient clearance of GAS in endothelial cells is caused by their intrinsic inability to target GAS with ubiquitin to promote autophagosome biogenesis for xenophagy. PMID:28683091

  4. Targeted Curing of All Lysogenic Bacteriophage from Streptococcus pyogenes Using a Novel Counter-selection Technique.

    PubMed

    Euler, Chad W; Juncosa, Barbara; Ryan, Patricia A; Deutsch, Douglas R; McShan, W Michael; Fischetti, Vincent A

    2016-01-01

    Streptococcus pyogenes is a human commensal and a bacterial pathogen responsible for a wide variety of human diseases differing in symptoms, severity, and tissue tropism. The completed genome sequences of >37 strains of S. pyogenes, representing diverse disease-causing serotypes, have been published. The greatest genetic variation among these strains is attributed to numerous integrated prophage and prophage-like elements, encoding several virulence factors. A comparison of isogenic strains, differing in prophage content, would reveal the effects of these elements on streptococcal pathogenesis. However, curing strains of prophage is often difficult and sometimes unattainable. We have applied a novel counter-selection approach to identify rare S. pyogenes mutants spontaneously cured of select prophage. To accomplish this, we first inserted a two-gene cassette containing a gene for kanamycin resistance (KanR) and the rpsL wild-type gene, responsible for dominant streptomycin sensitivity (SmS), into a targeted prophage on the chromosome of a streptomycin resistant (SmR) mutant of S. pyogenes strain SF370. We then applied antibiotic counter-selection for the re-establishment of the KanS/SmR phenotype to select for isolates cured of targeted prophage. This methodology allowed for the precise selection of spontaneous phage loss and restoration of the natural phage attB attachment sites for all four prophage-like elements in this S. pyogenes chromosome. Overall, 15 mutants were constructed that encompassed every permutation of phage knockout as well as a mutant strain, named CEM1ΔΦ, completely cured of all bacteriophage elements (a ~10% loss of the genome); the only reported S. pyogenes strain free of prophage-like elements. We compared CEM1ΔΦ to the WT strain by analyzing differences in secreted DNase activity, as well as lytic and lysogenic potential. These mutant strains should allow for the direct examination of bacteriophage relationships within S. pyogenes and

  5. Comparative growth, cross stress resistance, transcriptomics of Streptococcus pyogenes cultured under low shear modeled microgravity and normal gravity

    PubMed Central

    Kalpana, Duraisamy; Im, Chanki; Lee, Yang Soo

    2015-01-01

    Streptococcus pyogenes is commonly found on pharynx, mouth and rarely on skin, lower gastrointestinal tract. It is a potential pathogen causing tonsillitis, pneumonia, endocarditis. The present study was undertaken to study the effects of low shear modeled microgravity on growth, morphology, antibiotic resistance, cross-stress resistance to various stresses and alteration in gene expression of S. pyogenes. The growth analysis performed using UV–Visible spectroscopy indicated decrease in growth of S. pyogenes under low shear modeled microgravity. Morphological analysis by Bio-transmission electron microscopy (TEM), Bio-scanning electron microscopy (SEM) did not reveal much difference between normal and low shear modeled microgravity grown S. pyogenes. The sensitivity of S. pyogenes to antibiotics ampicillin, penicillin, streptomycin, kanamycin, hygromycin, rifampicin indicates that the bacterium is resistant to hygromycin. Further S. pyogenes cultured under low shear modeled microgravity was found to be more sensitive to ampicillin and rifampicin as compared with normal gravity grown S. pyogenes. The bacteria were tested for the acid, osmotic, temperature and oxidative cross stress resistances. The gene expression of S. pyogenes under low shear modeled microgravity analyzed by microarray revealed upregulation of 26 genes and down regulation of 22 genes by a fold change of 1.5. PMID:26858535

  6. Comparative growth, cross stress resistance, transcriptomics of Streptococcus pyogenes cultured under low shear modeled microgravity and normal gravity.

    PubMed

    Kalpana, Duraisamy; Im, Chanki; Lee, Yang Soo

    2016-01-01

    Streptococcus pyogenes is commonly found on pharynx, mouth and rarely on skin, lower gastrointestinal tract. It is a potential pathogen causing tonsillitis, pneumonia, endocarditis. The present study was undertaken to study the effects of low shear modeled microgravity on growth, morphology, antibiotic resistance, cross-stress resistance to various stresses and alteration in gene expression of S. pyogenes. The growth analysis performed using UV-Visible spectroscopy indicated decrease in growth of S. pyogenes under low shear modeled microgravity. Morphological analysis by Bio-transmission electron microscopy (TEM), Bio-scanning electron microscopy (SEM) did not reveal much difference between normal and low shear modeled microgravity grown S. pyogenes. The sensitivity of S. pyogenes to antibiotics ampicillin, penicillin, streptomycin, kanamycin, hygromycin, rifampicin indicates that the bacterium is resistant to hygromycin. Further S. pyogenes cultured under low shear modeled microgravity was found to be more sensitive to ampicillin and rifampicin as compared with normal gravity grown S. pyogenes. The bacteria were tested for the acid, osmotic, temperature and oxidative cross stress resistances. The gene expression of S. pyogenes under low shear modeled microgravity analyzed by microarray revealed upregulation of 26 genes and down regulation of 22 genes by a fold change of 1.5.

  7. Crystal structure of SP-PTP, a low molecular weight protein tyrosine phosphatase from Streptococcus pyogenes.

    PubMed

    Ku, Bonsu; Keum, Chae Won; Lee, Hye Seon; Yun, Hye-Yeoung; Shin, Ho-Chul; Kim, Bo Yeon; Kim, Seung Jun

    2016-09-23

    Streptococcus pyogenes, or Group A Streptococcus (GAS), is a pathogenic bacterium that causes a variety of infectious diseases. The GAS genome encodes one protein tyrosine phosphatase, SP-PTP, which plays an essential role in the replication and virulence maintenance of GAS. Herein, we present the crystal structure of SP-PTP at 1.9 Å resolution. Although SP-PTP has been reported to have dual phosphatase specificity for both phosphorylated tyrosine and serine/threonine, three-dimensional structural analysis showed that SP-PTP shares high similarity with typical low molecular weight protein tyrosine phosphatases (LMWPTPs), which are specific for phosphotyrosine, but not with dual-specificity phosphatases, in overall folding and active site composition. In the dephosphorylation activity test, SP-PTP consistently acted on phosphotyrosine substrates, but not or only minimally on phosphoserine/phosphothreonine substrates. Collectively, our structural and biochemical analyses verified SP-PTP as a canonical tyrosine-specific LMWPTP.

  8. Purification, crystallization and preliminary crystallographic analysis of Streptococcus pyogenes laminin-binding protein Lbp

    PubMed Central

    Linke, Christian; Caradoc-Davies, Tom T.; Proft, Thomas; Baker, Edward N.

    2008-01-01

    The laminin-binding protein Lbp (Spy2007) from Streptococcus pyogenes (a group A streptococcus) mediates adhesion to the human basal lamina glycoprotein laminin. Accordingly, Lbp is essential in in vitro models of cell adhesion and invasion. However, the molecular and structural basis of laminin binding by bacteria remains unknown. Therefore, the lbp gene has been cloned for recombinant expression in Escherichia coli. Lbp has been purified and crystallized from 30%(w/v) PEG 1500 by the sitting-drop vapour-diffusion method. The crystals belonged to the monoclinic space group P21, with unit-cell parameters a = 42.62, b = 92.16, c = 70.61 Å, β = 106.27°, and diffracted to 2.5 Å resolution. PMID:18259070

  9. Zinc disrupts central carbon metabolism and capsule biosynthesis in Streptococcus pyogenes

    PubMed Central

    Ong, Cheryl-lynn Y.; Walker, Mark J.; McEwan, Alastair G.

    2015-01-01

    Neutrophils release free zinc to eliminate the phagocytosed bacterial pathogen Streptococcus pyogenes (Group A Streptococcus; GAS). In this study, we investigated the mechanisms underpinning zinc toxicity towards this human pathogen, responsible for diseases ranging from pharyngitis and impetigo, to severe invasive infections. Using the globally-disseminated M1T1 GAS strain, we demonstrate that zinc stress impairs glucose metabolism through the inhibition of the glycolytic enzymes phosphofructokinase and glyceraldehyde-3-phosphate dehydrogenase. In the presence of zinc, a metabolic shift to the tagatose-6-phosphate pathway allows conversion of D-galactose to dihydroxyacetone phosphate and glyceraldehyde phosphate, partially bypassing impaired glycolytic enzymes to generate pyruvate. Additionally, zinc inhibition of phosphoglucomutase results in decreased capsule biosynthesis. These data indicate that zinc exerts it toxicity via mechanisms that inhibit both GAS central carbon metabolism and virulence pathways. PMID:26028191

  10. Streptococcus pyogenes Triggers Activation of the Human Contact System by Streptokinase

    PubMed Central

    Nitzsche, Ramona; Rosenheinrich, Maik; Kreikemeyer, Bernd

    2015-01-01

    Severe invasive infectious diseases remain a major and life-threatening health problem. In serious cases, a systemic activation of the coagulation cascade is a critical complication that is associated with high mortality rates. We report here that streptokinase, a group A streptococcal plasminogen activator, triggers the activation of the human contact system. Activation of contact system factors at the surface of the Streptococcus pyogenes serotype M49 is dependent on streptokinase and plasminogen. Our results also show that secreted streptokinase is an efficient contact system activator, independent from a contact surface. This results in the processing of high-molecular-weight kininogen and the release of bradykinin, a potent vascular mediator. We further investigated whether the ability of 50 different clinical S. pyogenes isolates to activate the contact system is associated with an invasive phenotype. The data reveal that isolates from invasive infections trigger an activation of the contact system more potently than strains isolated from noninvasive infections. The present study gives new insights into the mechanisms by which S. pyogenes triggers the human contact system and stresses the function of soluble and surface located plasmin exploited as a group A streptococcal virulence factor through the action of streptokinase. PMID:25987706

  11. Streptococcus pyogenes Pneumonia in Adults: Clinical Presentation and Molecular Characterization of Isolates 2006-2015

    PubMed Central

    Tamayo, Esther; Montes, Milagrosa; Vicente, Diego; Pérez-Trallero, Emilio

    2016-01-01

    Introduction In the preantibiotic era Streptococcus pyogenes was a common cause of severe pneumonia but currently, except for postinfluenza complications, it is not considered a common cause of community-acquired pneumonia in adults. Aim and Material and Methods This study aimed to identify current clinical episodes of S. pyogenes pneumonia, its relationship with influenza virus circulation and the genotypes of the involved isolates during a decade in a Southern European region (Gipuzkoa, northern Spain). Molecular analysis of isolates included emm, multilocus-sequence typing, and superantigen profile determination. Results Forty episodes were detected (annual incidence 1.1 x 100,000 inhabitants, range 0.29–2.29). Thirty-seven episodes were community-acquired, 21 involved an invasive infection and 10 developed STSS. The associated mortality rate was 20%, with half of the patients dying within 24 hours after admission. Influenza coinfection was confirmed in four patients and suspected in another. The 52.5% of episodes occurred outside the influenza seasonal epidemic. The 67.5% of affected persons were elderly individuals and adults with severe comorbidities, although 13 patients had no comorbidities, 2 of them had a fatal outcome. Eleven clones were identified, the most prevalent being emm1/ST28 (43.6%) causing the most severe cases. Conclusions S. pyogenes pneumonia had a continuous presence frequently unrelated to influenza infection, being rapidly fatal even in previously healthy individuals. PMID:27027618

  12. Macrolide resistance in Streptococcus pyogenes isolates from throat infections in the region of Aachen, Germany.

    PubMed

    Brandt, C M; Honscha, M; Truong, N D; Holland, R; Hövener, B; Bryskier, A; Lütticken, R; Reinert, R R

    2001-01-01

    Macrolide-resistance was assessed in 216 consecutive Streptococcus pyogenes isolates from throat infections in the region of Aachen, Germany. Seventeen isolates were resistant to erythromycin: 12 isolates revealed a macrolide (M) phenotype and harbored mefA, and five strains expressed an inducible macrolide-lincosamide-streptogramin B (MLSB) phenotype of which four strains harbored ermA(TR) and one strain contained ermB(AM). Telithromycin (HMR 3647) and quinupristin/dalfopristin remained active particularly against the ermA(TR)-containing S. pyogenes isolates studied. Random amplified polymorphic DNA analysis identified multiple clones among erythromycin-resistant strains, but did not discriminate beyond the emm-type. mefA was present in three isolates either with emm2, emm12, or emm75, and in nine isolates with emm4. All four strains with ermA(TR) contained emm77, and the single strain with ermB(AM) harbored emm1. Despite the relative low rate of macrolide-resistance, these data suggest that at least three different macrolide-resistance determinants are prevalent in Germany and that mefA has spread rapidly into multiple clones of S. pyogenes.

  13. Streptococcus pyogenes Arginine and Citrulline Catabolism Promotes Infection and Modulates Innate Immunity

    PubMed Central

    Cusumano, Zachary T.; Watson, Michael E.

    2014-01-01

    A bacterium's ability to acquire nutrients from its host during infection is an essential component of pathogenesis. For the Gram-positive pathogen Streptococcus pyogenes, catabolism of the amino acid arginine via the arginine deiminase (ADI) pathway supplements energy production and provides protection against acid stress in vitro. Its expression is enhanced in murine models of infection, suggesting an important role in vivo. To gain insight into the function of the ADI pathway in pathogenesis, the virulence of mutants defective in each of its enzymes was examined. Mutants unable to use arginine (ΔArcA) or citrulline (ΔArcB) were attenuated for carriage in a murine model of asymptomatic mucosal colonization. However, in a murine model of inflammatory infection of cutaneous tissue, the ΔArcA mutant was attenuated but the ΔArcB mutant was hyperattenuated, revealing an unexpected tissue-specific role for citrulline metabolism in pathogenesis. When mice defective for the arginine-dependent production of nitric oxide (iNOS−/−) were infected with the ΔArcA mutant, cutaneous virulence was rescued, demonstrating that the ability of S. pyogenes to utilize arginine was dispensable in the absence of nitric oxide-mediated innate immunity. This work demonstrates the importance of arginine and citrulline catabolism and suggests a novel mechanism of virulence by which S. pyogenes uses its metabolism to modulate innate immunity through depletion of an essential host nutrient. PMID:24144727

  14. [Invasive Streptococcus pyogenes diseases 2005-2013: Pediatric Hospital Pereira Rossell Uruguay].

    PubMed

    Vomero, Alejandra; García, Gabriela; Pandolfo, Soledad; Zunino, Carlos; Ambrosoni, María; Algorta, Gabriela; Pírez, M Catalina

    2014-12-01

    Streptococcus pyogenes infection causes a wide spectrum of clinical manifestations. Invasive disease (ID) is defined by the isolation of the microorganism from sterile sites. To analyze the clinical, epidemiological and molecular characteristics of ID by S.pyogenes in children hospitalized at Pediatric Hospital Pereira Rossell, from January 2005 to January 2013. A descriptive retrospective study was done in cases with isolation of S.pyogenes from sterile sites. Epidemiological variables, disease characteristics, laboratory parameters, treatment regimen received, hospitalization days and clinical outcome were analyzed. The presence of 4 genes encoding for virulence factors and chromosome profile studied by pulsed-field electrophoresis were done in the isolated strains. A total of 42 cases (rate: 4.6 out of 10,000 admissions) were detected, from which 32 were analyzed. Average age was 44.7 months (14/32 < 2 years of age). In 5 cases, the portal of entry was identified. Clinical presentations were: osteoarticular infections (n = 15), shock (n = 6), skin and soft-tissue infections (n = 5), pneumonia (n = 3) and bacteremias (n = 3). Twenty cases required surgical procedures and 13 required intensive care admission. Average hospital stay was 17 days and one patient died. Molecular studies were performed in five strains; 4 different toxin profiles and pulsotypes were identified. The incidence of ID at our hospital is similar to other series in the region. A better knowledge of clinical presentation and its relation with molecular characteristics represents a challenge.

  15. Epidemiology of necrotizing infection caused by Staphylococcus aureus and Streptococcus pyogenes at an Iowa hospital.

    PubMed

    Thapaliya, Dipendra; O'Brien, Ashley M; Wardyn, Shylo E; Smith, Tara C

    2015-01-01

    The present study was performed to characterize the epidemiology of necrotizing soft tissue infection caused by Streptococcus pyogenes (n=14) and Staphylococcus aureus (n=14) isolates collected at the University of Iowa Hospitals and Clinics. An additional 9 S. pyogenes isolates were collected from patients being treated for mild respiratory infections and served as a comparison sample in the analysis. Patient data corresponding to the isolates (n=37) were also collected in order to identify risk factors or comorbid conditions possibly correlated with necrotizing fasciitis (NF). The prevalence of methicillin-resistant S. aureus among the study isolates was 35.7% (5/14), and the prevalence of the Panton-Valentine leukocidin (PVL) gene was 57% (8/14). The S. pyogenes NF (wound) isolates (n=14) belonged to 10 different emm types, none of which appeared to be associated with more severe disease when compared to the milder infection (throat) samples (n=9). Comorbid conditions such as diabetes and cardiovascular disease were significantly associated with NF. The results indicate that there may be a high prevalence of the PVL virulence factor in NF infections and that spa type t008 may be responsible for the increasing incidence of S. aureus NF infections in Iowa.

  16. [Resistance phenotypes and genotypes of Streptococcus pyogenes clinical isolates in Chile over a 10-year period].

    PubMed

    Rodríguez, Carlos; Rojas, Pablo; Wozniak, Aniela; Kalergis, Alexis M; Cerón, Inés; Riedel, Ingrid; Román, Juan C; Villarroel, Luis A; Berríos, Ximena; Bavestrello, Luis; García, Patricia

    2011-09-01

    Macrolide and lincosamide resistance in Streptococcus pyogenes is due to the acquisition of mef, ermB and ermA genes, which confer different resistance phenotypes, namely M, MLSBconstitutive and MLSBinducible respectively. The last report of resistance in Chile was done in the period 1990-1998, in which resistance to macrolides was 5.4%, with M phenotype as the predominant one. To characterize the evolution of erythromycin and clindamycin resistance and their associated genes in S. pyogenes strains isolated from patients with invasive and non-invasive infections in the period 1996 to 2005. Resistance to erythromycin and clindamycin was determined in 1,282 clinical isolates using the disk diffusion test. Resistant isolates were analyzed by polymerase chain reaction (PCR) for the presence of the above mentioned resistance genes. Global resistance to erythromycin and clindamycin was 3.5 and 0.7% respectively. Eighty percent of the resistant strains possessed the M. phenotype. Resistance levels of S. pyogenes have decreased in Chile in the last years. Most resistant strains have M phenotype in contrast to many countries in which the MLSB constitutive phenotype is the predominant one.

  17. Two-Component Systems Involved in Susceptibility to Nisin A in Streptococcus pyogenes

    PubMed Central

    Kawada-Matsuo, Miki; Tatsuno, Ichiro; Arii, Kaoru; Zendo, Takeshi; Oogai, Yuichi; Noguchi, Kazuyuki; Hasegawa, Tadao; Sonomoto, Kenji

    2016-01-01

    ABSTRACT Two-component systems (TCSs) are regulatory systems in bacteria that play important roles in sensing and adapting to the environment. In this study, we systematically evaluated the roles of TCSs in the susceptibility of the group A Streptococcus (GAS; Streptococcus pyogenes) SF370 strain to several types of lantibiotics. Using individual TCS deletion mutants, we found that the deletion of srtRK (spy_1081–spy_1082) in SF370 increased the susceptibility to nisin A, which is produced by Lactococcus lactis ATCC 11454, but susceptibility to other types of lantibiotics (nukacin ISK-1, produced by Staphylococcus warneri, and staphylococcin C55, produced by Staphylococcus aureus) was not altered in the TCS mutants tested. The expression of srtFEG (spy_1085 to spy_1087), which is located downstream of srtRK and is homologous to ABC transporters, was increased in response to nisin A. However, srtEFG expression was not induced by nisin A in the srtRK mutant. The inactivation of srtFEG increased the susceptibility to nisin A. These results suggest that SrtRK controls SrtFEG expression to alter the susceptibility to nisin A. Further experiments showed that SrtRK is required for coexistence with L. lactis ATCC 11454, which produces nisin A. Our results elucidate the important roles of S. pyogenes TCSs in the interactions between different bacterial species, including bacteriocin-producing bacteria. IMPORTANCE In this study, we focused on the association of TCSs with susceptibility to bacteriocins in S. pyogenes SF370, which has no ability to produce bacteriocins, and reported two major new findings. We demonstrated that the SrtRK TCS is related to susceptibility to nisin A by controlling the ABC transporter SrtFEG. We also showed that S. pyogenes SrtRK is important for survival when the bacteria are cocultured with nisin A-producing Lactococcus lactis. This report highlights the roles of TCSs in the colocalization of bacteriocin-producing bacteria and non

  18. Lactobacilli Interfere with Streptococcus pyogenes Hemolytic Activity and Adherence to Host Epithelial Cells

    PubMed Central

    Saroj, Sunil D.; Maudsdotter, Lisa; Tavares, Raquel; Jonsson, Ann-Beth

    2016-01-01

    Streptococcus pyogenes [Group A streptococcus (GAS)], a frequent colonizer of the respiratory tract mucosal surface, causes a variety of human diseases, ranging from pharyngitis to the life-threatening streptococcal toxic shock-like syndrome. Lactobacilli have been demonstrated to colonize the respiratory tract. In this study, we investigated the interference of lactobacilli with the virulence phenotypes of GAS. The Lactobacillus strains L. rhamnosus Kx151A1 and L. reuteri PTA-5289, but not L. salivarius LMG9477, inhibited the hemolytic activity of S. pyogenes S165. The inhibition of hemolytic activity was attributed to a decrease in the production of streptolysin S (SLS). Conditioned medium (CM) from the growth of L. rhamnosus Kx151A1 and L. reuteri PTA-5289 was sufficient to down-regulate the expression of the sag operon, encoding SLS. The Lactobacillus strains L. rhamnosus Kx151A1, L. reuteri PTA-5289, and L. salivarius LMG9477 inhibited the initial adherence of GAS to host epithelial cells. Intriguingly, competition with a combination of Lactobacillus species reduced GAS adherence to host cells most efficiently. The data suggest that an effector molecule released from certain Lactobacillus strains attenuates the production of SLS at the transcriptional level and that combinations of Lactobacillus strains may protect the pharyngeal mucosa more efficiently from the initial colonization of GAS. The effector molecules released from Lactobacillus strains affecting the virulence phenotypes of pathogens hold potential in the development of a new generation of therapeutics. PMID:27524981

  19. Identification of Streptococcus pyogenes – Phenotypic Tests vs Molecular Assay (spy1258PCR): A Comparative Study

    PubMed Central

    Abraham, Tintu

    2016-01-01

    Introduction Traditionally Group A Streptococcus pyogenes (GAS) is differentiated from other beta haemolytic streptococci (BHS) by certain presumptive tests such as bacitracin sensitivity and production of Pyrollidonyl Aryl Sulfatase (PYR). The phenotypic and genotypic confirmatory tests are Lancefield grouping for cell wall carbohydrate antigen and PCR for spy1258 gene respectively. Reliance on presumptive tests alone may lead to misidentification of isolates. Aim To compare the predictive values of routine phenotypic tests with spy1258 PCR for the identification of Streptococcus pyogenes. Materials and Methods This comparative analytical study was carried out in the Department of Microbiology, JIPMER, Puducherry, over a period of 18 months (1st November 2013 to 30th April 2015). Two hundred and six consecutive BHS isolates from various clinical samples were subjected to phenotypic tests such as bacitracin sensitivity, PYR test and Lancefield grouping. The results were compared with spy1258 PCR which was considered 95 the confirmatory test for identification. Results The sensitivity and specificity of phenotypic tests were as follows; Susceptibility to bacitracin – 95.42%, 70.96%, PYR test – 95.42%, 77.41%, Lancefield grouping- 97.71%, 80.64%. Conclusion Clinical laboratories should not depend on bacitracin sensitivity as a single presumptive test for the routine identification of GAS but should use supplemental tests such as PYR test or latex agglutination test and for best results use spy1258 PCR. PMID:27630838

  20. Identification of Streptococcus pyogenes - Phenotypic Tests vs Molecular Assay (spy1258PCR): A Comparative Study.

    PubMed

    Abraham, Tintu; Sistla, Sujatha

    2016-07-01

    Traditionally Group A Streptococcus pyogenes (GAS) is differentiated from other beta haemolytic streptococci (BHS) by certain presumptive tests such as bacitracin sensitivity and production of Pyrollidonyl Aryl Sulfatase (PYR). The phenotypic and genotypic confirmatory tests are Lancefield grouping for cell wall carbohydrate antigen and PCR for spy1258 gene respectively. Reliance on presumptive tests alone may lead to misidentification of isolates. To compare the predictive values of routine phenotypic tests with spy1258 PCR for the identification of Streptococcus pyogenes. This comparative analytical study was carried out in the Department of Microbiology, JIPMER, Puducherry, over a period of 18 months (1(st) November 2013 to 30(th) April 2015). Two hundred and six consecutive BHS isolates from various clinical samples were subjected to phenotypic tests such as bacitracin sensitivity, PYR test and Lancefield grouping. The results were compared with spy1258 PCR which was considered 95 the confirmatory test for identification. The sensitivity and specificity of phenotypic tests were as follows; Susceptibility to bacitracin - 95.42%, 70.96%, PYR test - 95.42%, 77.41%, Lancefield grouping- 97.71%, 80.64%. Clinical laboratories should not depend on bacitracin sensitivity as a single presumptive test for the routine identification of GAS but should use supplemental tests such as PYR test or latex agglutination test and for best results use spy1258 PCR.

  1. Complete Genome Sequence of emm4 Streptococcus pyogenes MEW427, a Throat Isolate from a Child Meeting Clinical Criteria for Pediatric Autoimmune Neuropsychiatric Disorders Associated with Streptococcus (PANDAS).

    PubMed

    Jacob, Kristin M; Spilker, Theodore; LiPuma, John J; Dawid, Suzanne R; Watson, Michael E

    2016-03-17

    We report the complete genome assembly of the Streptococcus pyogenes type emm4 strain MEW427 (also referred to as strain UM001 in the Pediatric Acute-Onset Neuropsychiatric Syndrome [PANS] Research Consortium), a throat isolate from a child with acute-onset neuropsychiatric symptoms meeting clinical criteria for PANDAS (pediatric autoimmune neuropsychiatric disorders associated with streptococcus). The genome length is 1,814,455 bp with 38.51% G+C%. Copyright © 2016 Jacob et al.

  2. High macrolide resistance in Streptococcus pyogenes strains isolated from children with pharyngitis in China.

    PubMed

    Liu, Xiaorong; Shen, Xuzhuang; Chang, Hesheng; Huang, Guoying; Fu, Zhou; Zheng, Yuejie; Wang, Libo; Li, Chengrong; Liu, Lan; Shen, Ying; Yang, Yonghong

    2009-05-01

    To assess the macrolide resistance, phenotype, and genotypic characterization of Streptococcus pyogenes isolated from Chinese children with pharyngitis. Minimal inhibitory concentration (MIC) with nine antibiotics was determined on 188 isolates of S. pyogenes collected from outpatients with pharyngitis in four children's hospitals in different regions of China in 2007. MICs of penicillin, chloramphenicol, cefradine, levofloxacin, macrolide (erythromycin, clarithromycin, azithromycin,), clindamycin, and tetracycline were determined by the microdilution method. The macrolide resistant phenotypes of isolates were determined through a double-disk. The macrolide-resistant genes (mefA, ermB, and ermA) were amplified by polymerase chain reaction (PCR). Over 95% were resistant to macrolides, while 92.0% were resistant to tetracycline. We also found that all isolates were sensitive to penicillin, chloramphenicol, cefradine, and levofloxacin. Among the 173 erythromycin resistant strains, 171 (98.8%) were assigned to the cMLS phenotype, while the remaining 2 (1.2%) were assigned to the iMLS phenotype. Among the 171 cMLS isolates, 168 isolates (98.2%) had the ermB gene accounting for 98.2%. Meanwhile, 2 iMLS isolates had the ermA gene. Macrolides were highly resistant to ermB positive strains (MIC(90) > 256 microg/ml). Neither the M-phenotype nor the mefA gene was detected. Meanwhile, our studies of multiple centers showed that consumption of macrolides from 2000 to 2006 was very high. The main phenotype is cMLS, and the ermB gene code is the main resistance mechanism against macrolides in S. pyogenes. The high rate of macrolide resistance to S. pyogenes was observed, which may be correlated with the overuse of these antibiotics in China. (c) 2009 Wiley-Liss, Inc.

  3. Cysteine Proteinase from Streptococcus pyogenes Enables Evasion of Innate Immunity via Degradation of Complement Factors*

    PubMed Central

    Honda-Ogawa, Mariko; Ogawa, Taiji; Terao, Yutaka; Sumitomo, Tomoko; Nakata, Masanobu; Ikebe, Kazunori; Maeda, Yoshinobu; Kawabata, Shigetada

    2013-01-01

    Streptococcus pyogenes is an important human pathogen that causes invasive diseases such as necrotizing fasciitis, sepsis, and streptococcal toxic shock syndrome. We investigated the function of a major cysteine protease from S. pyogenes that affects the amount of C1-esterase inhibitor (C1-INH) and other complement factors and aimed to elucidate the mechanism involved in occurrence of streptococcal toxic shock syndrome from the aspect of the complement system. First, we revealed that culture supernatant of a given S. pyogenes strain and recombinant SpeB degraded the C1-INH. Then, we determined the N-terminal sequence of the C1-INH fragment degraded by recombinant SpeB. Interestingly, the region containing one of the identified cleavage sites is not present in patients with C1-INH deficiency. Scanning electron microscopy of the speB mutant incubated in human serum showed the abnormal superficial architecture and irregular oval structure. Furthermore, unlike the wild-type strain, that mutant strain showed lower survival capacity than normal as compared with heat-inactivated serum, whereas it had a significantly higher survival rate in serum without the C1-INH than in normal serum. Also, SpeB degraded multiple complement factors and the membrane attack complex. Flow cytometric analyses revealed deposition of C9, one of the components of membrane the attack complex, in greater amounts on the surface of the speB mutant, whereas lower amounts of C9 were bound to the wild-type strain surface. These results suggest that SpeB can interrupt the human complement system via degrading the C1-INH, thus enabling S. pyogenes to evade eradication in a hostile environment. PMID:23589297

  4. Telithromycin and quinupristin-dalfopristin resistance in clinical isolates of Streptococcus pyogenes: SMART Program 2001 Data.

    PubMed

    Hsueh, Po-Ren; Teng, Lee-Jene; Lee, Chun-Ming; Huang, Wen-Kuei; Wu, Tsu-Lan; Wan, Jen-Hsien; Yang, Dine; Shyr, Jainn-Ming; Chuang, Yin-Ching; Yan, Jing-Jou; Lu, Jang-Jih; Wu, Jiunn-Jong; Ko, Wen-Chien; Chang, Feng-Yee; Yang, Yi-Chueh; Lau, Yeu-Jun; Liu, Yung-Ching; Leu, Hsieh-Shong; Liu, Cheng-Yi; Luh, Kwen-Tay

    2003-07-01

    This study evaluated the current status of antimicrobial resistance in clinical isolates of Streptococcus pyogenes in Taiwan as part of the SMART (Surveillance from Multicenter Antimicrobial Resistance in Taiwan) program. In 2001, 419 different isolates of S. pyogenes, including 275 from respiratory secretions, 87 from wound pus, and 31 from blood, were collected from nine hospitals in different parts of Taiwan. MICs of 23 antimicrobial agents were determined at a central location by the agar dilution method. All of the isolates were susceptible to penicillin (MIC at which 90% of the isolates were inhibited [MIC(90)], moxifloxacin > ciprofloxacin = levofloxacin = gatifloxacin > gemifloxacin) demonstrated potent activity against nearly all of the isolates of S. pyogenes tested. Thirty-two isolates (8%) were not susceptible to quinupristin-dalfopristin. Seventeen percent of isolates had telithromycin MICs of >or=1 microg/ml, and all of these isolates exhibited erythromycin MICs of >or=32 microg/ml. The high prevalence of resistance to telithromycin (which is not available in Taiwan) limits its potential use in the treatment of S. pyogenes infections, particularly in areas with high rates of macrolide resistance.

  5. Long-term survival of Streptococcus pyogenes in rich media is pH-dependent.

    PubMed

    Savic, Dragutin J; McShan, William M

    2012-06-01

    The mechanisms that allow Streptococcus pyogenes to survive and persist in the human host, often in spite of antibiotic therapy, remain poorly characterized. Therefore, the determination of culture conditions for long-term studies is crucial to advancement in this field. Stationary cultures of S. pyogenes strain NZ131 and its spontaneous small-colony variant OK171 were found to survive in rich medium for less than 2 weeks, and this inability to survive resulted from the acidification of the medium to below pH 5.5, which the cells did not tolerate for longer than 6-7 days. The growth of NZ131 resulted in acidification of the culture to below pH 5.5 by the onset of stationary phase, and the loss of viability occurred in a linear fashion. These results were also found to be true for M49 strain CS101 and for M1 strain SF370. The S. pyogenes strains could be protected from killing by the addition of a buffer that stabilized the pH of the medium at pH 6.5, ensuring bacterial survival to at least 70 days. By contrast, increasing the glucose added to the medium accelerated the loss of culture viability in strain NZ131 but not OK171, suggesting that the small-colony variant is altered in glucose uptake or metabolism. Similarly, acidification of the medium prior to inoculation or at the middle of exponential phase resulted in growth inhibition of all strains. These results suggest that control of the pH is crucial for establishing long-term cultures of S. pyogenes.

  6. Cysteine proteinase from Streptococcus pyogenes enables evasion of innate immunity via degradation of complement factors.

    PubMed

    Honda-Ogawa, Mariko; Ogawa, Taiji; Terao, Yutaka; Sumitomo, Tomoko; Nakata, Masanobu; Ikebe, Kazunori; Maeda, Yoshinobu; Kawabata, Shigetada

    2013-05-31

    Streptococcus pyogenes is an important human pathogen that causes invasive diseases such as necrotizing fasciitis, sepsis, and streptococcal toxic shock syndrome. We investigated the function of a major cysteine protease from S. pyogenes that affects the amount of C1-esterase inhibitor (C1-INH) and other complement factors and aimed to elucidate the mechanism involved in occurrence of streptococcal toxic shock syndrome from the aspect of the complement system. First, we revealed that culture supernatant of a given S. pyogenes strain and recombinant SpeB degraded the C1-INH. Then, we determined the N-terminal sequence of the C1-INH fragment degraded by recombinant SpeB. Interestingly, the region containing one of the identified cleavage sites is not present in patients with C1-INH deficiency. Scanning electron microscopy of the speB mutant incubated in human serum showed the abnormal superficial architecture and irregular oval structure. Furthermore, unlike the wild-type strain, that mutant strain showed lower survival capacity than normal as compared with heat-inactivated serum, whereas it had a significantly higher survival rate in serum without the C1-INH than in normal serum. Also, SpeB degraded multiple complement factors and the membrane attack complex. Flow cytometric analyses revealed deposition of C9, one of the components of membrane the attack complex, in greater amounts on the surface of the speB mutant, whereas lower amounts of C9 were bound to the wild-type strain surface. These results suggest that SpeB can interrupt the human complement system via degrading the C1-INH, thus enabling S. pyogenes to evade eradication in a hostile environment.

  7. Importance of adhesins in the recurrence of pharyngeal infections caused by Streptococcus pyogenes.

    PubMed

    Wozniak, Aniela; Scioscia, Natalia; Geoffroy, Enrique; Ponce, Iván; García, Patricia

    2017-04-01

    Pharyngo-amygdalitis is the most common infection caused by Streptococcus pyogenes (S. pyogenes). Reinfection with strains of different M types commonly occurs. However, a second infection with a strain of the same M type can still occur and is referred to as recurrence. We aimed to assess whether recurrence of S. pyogenes could be associated to erythromycin resistance, biofilm formation or surface adhesins like fibronectin-binding proteins and pilus proteins, both located in the fibronectin-binding, collagen-binding, T-antigen (FCT) region. We analyed clinical isolates of S. pyogenes obtained from children with multiple positive cultures of throat swabs. We analysed potential associations between M types, clonal patterns, biofilm production and FCT types with their capacity of producing a recurrent infection. We genetically defined recurrence as an infection with the same M type (same strain) and reinfection as an infection with a different M type. No differences were observed between recurrent and reinfection isolates in relation to erythromycin resistance, presence and number of domains of prtF1 gene, and biofilm formation capacity; the only significant difference was the higher frequency of FCT-4 type among recurrent isolates. However, when all the factors that could contribute to recurrence (erythromycin resistance, biofilm production, presence of prtF1 gene and FCT-4 type) were analysed together, we observed that recurrent isolates have a higher number of factors than reinfection isolates. Recurrence seems not to be associated with biofilm formation. However, pili and fibronectin-binding proteins could be associated with recurrence because FCT-4 isolates which harbour two fibronectin-binding proteins are more frequent among recurrent isolates.

  8. Development of a high-throughput opsonophagocytic assay for the determination of functional antibody activity against Streptococcus pyogenes using bioluminescence.

    PubMed

    Lorenz, Natalie; Loh, Jacelyn M S; Moreland, Nicole J; Proft, Thomas

    2017-03-01

    The lack of standardised protocols for the assessment of functional antibodies has hindered Streptococcus pyogenes research and the development of vaccines. A robust, high throughput opsonophagocytic bactericidal assay to determine protective antibodies in human and rabbit serum has been developed that utilises bioluminescence as a rapid read out.

  9. Draft Genome Sequences of Two Streptococcus pyogenes Strains Involved in Abnormal Sharp Raised Scarlet Fever in China, 2011

    PubMed Central

    You, Yuanhai; Yang, Xianwei; Song, Yanyan; Yan, Xiaomei; Yuan, Yanting; Li, Dongfang; Yan, Yanfeng; Wang, Haibin; Tao, Xiaoxia; Li, Leilei; Jiang, Xihong; Zhou, Hao; Xiao, Di; Jin, Lianmei; Feng, Zijian; Yang, Ruifu; Luo, Fengji

    2012-01-01

    A scarlet fever outbreak caused by Streptococcus pyogenes occurred in China in 2011. To determine the genomic features of the outbreak strains, we deciphered genomes of two strains isolated from the regions with the highest incidence rates. The sequences will provide valuable information for comprehensive study of mechanisms related to this outbreak. PMID:23045496

  10. Characterization of macrolide efflux pump mef subclasses detected in clinical isolates of Streptococcus pyogenes isolated between 1999 and 2005.

    PubMed

    Blackman Northwood, J; Del Grosso, M; Cossins, L R; Coley, M D; Creti, R; Pantosti, A; Farrell, D J

    2009-05-01

    The macrolide efflux mechanism of resistance, mef, was characterized in community-acquired respiratory tract infections with Streptococcus pyogenes. Fifty-four (4.6%) M phenotype isolates were screen tested as negative for mef(A). Of these 54 isolates, 5 (0.4%), 27 (2.3%), and 1 (0.1%) were considered to be mef(I) positive, a novel mosaic variant of mef, or a novel subclass of mef, respectively. This study shows (i) the definitive presence of mef(E) in S. pyogenes and its global distribution, (ii) the presence of a mosaic variant of mef composed of mef(A) and mef(E), (iii) the previously undescribed presence of mef(I) in S. pyogenes, and (iv) the presence of a novel subclass of mef in S. pyogenes.

  11. Characterization of Macrolide Efflux Pump mef Subclasses Detected in Clinical Isolates of Streptococcus pyogenes Isolated between 1999 and 2005▿

    PubMed Central

    Blackman Northwood, J.; Del Grosso, M.; Cossins, L. R.; Coley, M. D.; Creti, R.; Pantosti, A.; Farrell, D. J.

    2009-01-01

    The macrolide efflux mechanism of resistance, mef, was characterized in community-acquired respiratory tract infections with Streptococcus pyogenes. Fifty-four (4.6%) M phenotype isolates were screen tested as negative for mef(A). Of these 54 isolates, 5 (0.4%), 27 (2.3%), and 1 (0.1%) were considered to be mef(I) positive, a novel mosaic variant of mef, or a novel subclass of mef, respectively. This study shows (i) the definitive presence of mef(E) in S. pyogenes and its global distribution, (ii) the presence of a mosaic variant of mef composed of mef(A) and mef(E), (iii) the previously undescribed presence of mef(I) in S. pyogenes, and (iv) the presence of a novel subclass of mef in S. pyogenes. PMID:19258262

  12. Prevention of streptococcal pharyngitis by anti-Streptococcus pyogenes bacteriocin-like inhibitory substances (BLIS) produced by Streptococcus salivarius.

    PubMed

    Tagg, J R

    2004-05-01

    Streptococcus salivarius is a numerically prominent member of the human oral microbiota that produces a variety of bacteriocin-like inhibitory substances (BLIS) having in vitro inhibitory activity against S. pyogenes. Our previous studies of S. salivarius isolates from children using a deferred antagonism BLIS production (P)-typing scheme showed that the 9 per cent of children having large populations of P-type 677 S. salivarius experienced fewer S. pyogenes acquisitions than either the 11 per cent of children having predominant P-type 226 populations or the 60 per cent of children with largely non-inhibitory (P-type 000) S. salivarius. Amongst the other BLIS P-types detected were a number of strongly-inhibitory (P-type 777) S. salivarius. In the present study the inhibitory agents produced by prototype strains of P-types 226, 677 and 777 S. salivarius are compared. The prototype BLIS-producing S. salivarius strains SN, 20P3, and K12 were isolated from tongue swabbings. BLIS P-typing was done using standard procedures. The BLIS molecules were purified and characterized. S. salivarius SN (P-type 226) produces a heat-labile muramidase. S. salivarius 20P3 (P-type 677) produces the 2315 Da lantibiotic salivaricin A and S. salivarius K12 (P-type 777) produces two lantibiotics; salivaricin A2 (2368 Da) and salivaricin B (2733 Da). The P-type 777 S. salivarius strain produced salivaricin A2 and salivaricin B. The combined production of two anti-S. pyogenes BLIS activities by this strain indicates that it could be adopted as a colonizing strain in bacterial interference trials.

  13. Frequency of Spontaneous Resistance to Peptide Deformylase Inhibitor GSK1322322 in Haemophilus influenzae, Staphylococcus aureus, Streptococcus pyogenes, and Streptococcus pneumoniae.

    PubMed

    Min, Sharon; Ingraham, Karen; Huang, Jianzhong; McCloskey, Lynn; Rilling, Sarah; Windau, Anne; Pizzollo, Jason; Butler, Deborah; Aubart, Kelly; Miller, Linda A; Zalacain, Magdalena; Holmes, David J; O'Dwyer, Karen

    2015-08-01

    The continuous emergence of multidrug-resistant pathogenic bacteria is compromising the successful treatment of serious microbial infections. GSK1322322, a novel peptide deformylase (PDF) inhibitor, shows good in vitro antibacterial activity and has demonstrated safety and efficacy in human proof-of-concept clinical studies. In vitro studies were performed to determine the frequency of resistance (FoR) to this antimicrobial agent in major pathogens that cause respiratory tract and skin infections. Resistance to GSK1322322 occurred at high frequency through loss-of-function mutations in the formyl-methionyl transferase (FMT) protein in Staphylococcus aureus (4/4 strains) and Streptococcus pyogenes (4/4 strains) and via missense mutations in Streptococcus pneumoniae (6/21 strains), but the mutations were associated with severe in vitro and/or in vivo fitness costs. The overall FoR to GSK1322322 was very low in Haemophilus influenzae, with only one PDF mutant being identified in one of four strains. No target-based mutants were identified from S. pyogenes, and only one or no PDF mutants were isolated in three of the four S. aureus strains studied. In S. pneumoniae, PDF mutants were isolated from only six of 21 strains tested; an additional 10 strains did not yield colonies on GSK1322322-containing plates. Most of the PDF mutants characterized from those three organisms (35/37 mutants) carried mutations in residues at or in close proximity to one of three highly conserved motifs that are part of the active site of the PDF protein, with 30 of the 35 mutations occurring at position V71 (using the S. pneumoniae numbering system). Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  14. The Transcriptional Regulator CpsY Is Important for Innate Immune Evasion in Streptococcus pyogenes

    PubMed Central

    Vega, Luis A.; Valdes, Kayla M.; Sundar, Ganesh S.; Belew, Ashton T.; Islam, Emrul; Berge, Jacob; Curry, Patrick; Chen, Steven

    2016-01-01

    ABSTRACT As an exclusively human pathogen, Streptococcus pyogenes (the group A streptococcus [GAS]) has specifically adapted to evade host innate immunity and survive in multiple tissue niches, including blood. GAS can overcome the metabolic constraints of the blood environment and expresses various immunomodulatory factors necessary for survival and immune cell resistance. Here we present our investigation of one such factor, the predicted LysR family transcriptional regulator CpsY. The encoding gene, cpsY, was initially identified as being required for GAS survival in a transposon-site hybridization (TraSH) screen in whole human blood. CpsY is homologous with transcriptional regulators of Streptococcus mutans (MetR), Streptococcus iniae (CpsY), and Streptococcus agalactiae (MtaR) that regulate methionine transport, amino acid metabolism, resistance to neutrophil-mediated killing, and survival in vivo. Our investigation indicated that CpsY is involved in GAS resistance to innate immune cells of its human host. However, GAS CpsY does not manifest the in vitro phenotypes of its homologs in other streptococcal species. GAS CpsY appears to regulate a small set of genes that is markedly different from the regulons of its homologs. The differential expression of these genes depends on the growth medium, and CpsY modestly influences their expression. The GAS CpsY regulon includes known virulence factors (mntE, speB, spd, nga [spn], prtS [SpyCEP], and sse) and cell surface-associated factors of GAS (emm1, mur1.2, sibA [cdhA], and M5005_Spy0500). Intriguingly, the loss of CpsY in GAS does not result in virulence defects in murine models of infection, suggesting that CpsY function in immune evasion is specific to the human host. PMID:27993974

  15. Salivaricin G32, a Homolog of the Prototype Streptococcus pyogenes Nisin-Like Lantibiotic SA-FF22, Produced by the Commensal Species Streptococcus salivarius.

    PubMed

    Wescombe, Philip A; Dyet, Kristin H; Dierksen, Karen P; Power, Daniel A; Jack, Ralph W; Burton, Jeremy P; Inglis, Megan A; Wescombe, Anna L; Tagg, John R

    2012-01-01

    Salivaricin G32, a 2667 Da novel member of the SA-FF22 cluster of lantibiotics, has been purified and characterized from Streptococcus salivarius strain G32. The inhibitory peptide differs from the Streptococcus pyogenes-produced SA-FF22 in the absence of lysine in position 2. The salivaricin G32 locus was widely distributed in BLIS-producing S. salivarius, with 6 (23%) of 26 strains PCR-positive for the structural gene, slnA. As for most other lantibiotics produced by S. salivarius, the salivaricin G32 locus can be megaplasmid encoded. Another member of the SA-FF22 family was detected in two Streptococcus dysgalactiae of bovine origin, an observation supportive of widespread distribution of this lantibiotic within the genus Streptococcus. Since the inhibitory spectrum of salivaricin G32 includes Streptococcus pyogenes, its production by S. salivarius, either as a member of the normal oral microflora or as a commercial probiotic, could serve to enhance protection of the human host against S. pyogenes infection.

  16. Geoepidemiological hints about Streptococcus pyogenes strains in relationship with acute rheumatic fever.

    PubMed

    Esposito, Susanna; Bianchini, Sonia; Fastiggi, Michele; Fumagalli, Monica; Andreozzi, Laura; Rigante, Donato

    2015-07-01

    Group A Streptococcus (GAS) strains are lately classified on the basis of sequence variations in the emm gene encoding the M protein, but despite the high number of distinct emm genotypes, the spectrum of phenotypes varying from invasive suppurative to non-suppurative GAS-related disorders has still to be defined. The relationship of GAS types with the uprising of acute rheumatic fever (ARF), a multisystemic disease caused by misdirected anti-GAS response in predisposed people, is also obscure. Studies published over the last 15 years were retrieved from PubMed using the keywords: "Streptococcus pyogenes" or "group A Streptococcus" and "acute rheumatic fever": the prevalence of peculiar emm types across different countries of the world is highly variable, depending on research designs, year of observation, country involved, patients' age, and gender. Most studies revealed that a relatively small number of specific emm/M protein types can be considered "rheumatogenic", as potentially characterized by the possibility of inducing ARF, with remarkable differences between developing and developed countries. The association between emm types and post-streptococcal manifestations is challenging, however surveillance of disease-causing variants in a specific community with high rate of ARF should be reinforced with the final goal of developing a potential primary prophylaxis against GAS infections.

  17. Manuka honey inhibits the development of Streptococcus pyogenes biofilms and causes reduced expression of two fibronectin binding proteins.

    PubMed

    Maddocks, Sarah E; Lopez, Marta Salinas; Rowlands, Richard S; Cooper, Rose A

    2012-03-01

    Streptococcus pyogenes (group A Streptococcus; GAS) is always of clinical significance in wounds where it can initiate infection, destroy skin grafts and persist as a biofilm. Manuka honey has broad spectrum antimicrobial activity and its use in the clinical setting is beginning to gain acceptance with the continuing emergence of antibiotic resistance and the inadequacy of established systemic therapies; novel inhibitors may affect clinical practice. In this study, the effect of manuka honey on S. pyogenes (M28) was investigated in vitro with planktonic and biofilm cultures using MIC, MBC, microscopy and aggregation efficiency. Bactericidal effects were found in both planktonic cultures and biofilms, although higher concentrations of manuka honey were needed to inhibit biofilms. Abrogation of adherence and intercellular aggregation was observed. Manuka honey permeated 24 h established biofilms of S. pyogenes, resulting in significant cell death and dissociation of cells from the biofilm. Sublethal concentrations of manuka honey effectively prevented the binding of S. pyogenes to the human tissue protein fibronectin, but did not inhibit binding to fibrinogen. The observed inhibition of fibronectin binding was confirmed by a reduction in the expression of genes encoding two major fibronectin-binding streptococcal surface proteins, Sof and SfbI. These findings indicate that manuka honey has potential in the topical treatment of wounds containing S. pyogenes.

  18. Variation in Streptococcus pyogenes NAD+ glycohydrolase is associated with tissue tropism.

    PubMed

    Riddle, David J; Bessen, Debra E; Caparon, Michael G

    2010-07-01

    Streptococcus pyogenes is an important pathogen that causes a variety of diseases. The most common infections involve the throat (pharyngitis) or skin (impetigo); however, the factors that determine tissue tropism and severity are incompletely understood. The S. pyogenes NAD(+) glycohydrolase (SPN) is a virulence factor that has been implicated in contributing to the pathogenesis of severe infections. However, the role of SPN in determining the bacterium's tissue tropism has not been evaluated. In this report, we examine the sequences of spn and its endogenous inhibitor ifs from a worldwide collection of S. pyogenes strains. Analysis of average pairwise nucleotide diversity, average number of nucleotide differences, and ratio of nonsynonymous to synonymous substitutions revealed significant diversity in spn and ifs. Application of established models of molecular evolution shows that SPN is evolving under positive selection and diverging into NAD(+) glycohydrolase (NADase)-active and -inactive subtypes. Additionally, the NADase-inactive SPN subtypes maintain the characteristics of a functional gene while ifs becomes a pseudogene. Thus, NADase-inactive SPN continues to evolve under functional constraint. Furthermore, NADase activity did not correlate with invasive disease in our collection but was associated with tissue tropism. The ability to cause infection at both the pharynx and the skin ("generalist" strains) is correlated with NADase-active SPN, while the preference for causing infection at either the throat or the skin ("specialist" strains) is associated with NADase-inactive SPN. These findings suggest that SPN has a NADase-independent function and prompt a reevaluation of the role of SPN in streptococcal pathogenesis.

  19. HMGB1 in severe soft tissue infections caused by Streptococcus pyogenes.

    PubMed

    Johansson, Linda; Snäll, Johanna; Sendi, Parham; Linnér, Anna; Thulin, Pontus; Linder, Adam; Treutiger, Carl-Johan; Norrby-Teglund, Anna

    2014-01-01

    Extracellular High Mobility Group Box 1 (HMGB1) has been associated with acute and chronic inflammatory conditions. However, little is known about HMGB1 in necrotizing bacterial infections. We hypothesized that the local HMGB1 response is excessive in severe soft tissue infections (STIs), which are characterized by necrosis and hyperinflammation. To explore this, tissue biopsies were collected from patients with varying severity of Streptococcus pyogenes skin and STIs, including erysipelas, cellulitis, and necrotizing fasciitis. Tissue sections were immunostained for HMGB1, S. pyogenes, and inflammatory cell infiltrates and results quantified by acquired computerized image analysis (ACIA). HMGB1 expression increased in parallel to disease severity and was significantly higher in necrotizing fasciitis than in erysipelas (p = 0.0023). Confocal microscopy of sections co-stained for HMGB1 and cell markers revealed both extracellular and cytoplasmic HMGB1, the latter of which was found predominantly in macrophages. To further verify macrophages as main source of activation triggered HMGB1 release, human macrophages were infected with clinical S. pyogenes isolates. The results demonstrated infection triggered release of HMGB1. Dual staining's visualized HMGB1 in areas close to, but not overlapping, with neutrophils, indicating a potential chemotactic role. In vitro transmigration experiments showed a chemotactic effect of HMGB1 on neutrophils. The data furthermore provided in vivo support that HGMB1 may form immunostimulatory complexes with IL-1β. Taken together, the findings provide the first in vivo evidence that HMGB1 is abundant at the local site of severe bacterial STIs and its levels correlated to severity of infections; hence, indicating its potential value as a biomarker for tissue pathology.

  20. Streptococcus pyogenes Malate Degradation Pathway Links pH Regulation and Virulence

    PubMed Central

    Paluscio, Elyse

    2015-01-01

    The ability of Streptococcus pyogenes to infect different niches within its human host most likely relies on its ability to utilize alternative carbon sources. In examining this question, we discovered that all sequenced S. pyogenes strains possess the genes for the malic enzyme (ME) pathway, which allows malate to be used as a supplemental carbon source for growth. ME is comprised of four genes in two adjacent operons, with the regulatory two-component MaeKR required for expression of genes encoding a malate permease (maeP) and malic enzyme (maeE). Analysis of transcription indicated that expression of maeP and maeE is induced by both malate and low pH, and induction in response to both cues is dependent on the MaeK sensor kinase. Furthermore, both maePE and maeKR are repressed by glucose, which occurs via a CcpA-independent mechanism. Additionally, malate utilization requires the PTS transporter EI enzyme (PtsI), as a PtsI– mutant fails to express the ME genes and is unable to utilize malate. Virulence of selected ME mutants was assessed in a murine model of soft tissue infection. MaeP–, MaeK–, and MaeR– mutants were attenuated for virulence, whereas a MaeE– mutant showed enhanced virulence compared to that of the wild type. Taken together, these data show that ME contributes to S. pyogenes' carbon source repertory, that malate utilization is a highly regulated process, and that a single regulator controls ME expression in response to diverse signals. Furthermore, malate uptake and utilization contribute to the adaptive pH response, and ME can influence the outcome of infection. PMID:25583521

  1. Aberrant Inflammatory Response to Streptococcus pyogenes in Mice Lacking Myeloid Differentiation Factor 88

    PubMed Central

    Loof, Torsten G.; Goldmann, Oliver; Gessner, André; Herwald, Heiko; Medina, Eva

    2010-01-01

    Several in vitro studies have emphasized the importance of toll-like receptor/myeloid differentiation factor 88 (MyD88) signaling in the inflammatory response to Streptococcus pyogenes. Since the extent of inflammation has been implicated in the severity of streptococcal diseases, we have examined here the role of toll-like receptor/MyD88 signaling in the pathophysiology of experimental S. pyogenes infection. To this end, we compared the response of MyD88-knockout (MyD88−/−) after subcutaneous inoculation with S. pyogenes with that of C57BL/6 mice. Our results show that MyD88−/− mice harbored significantly more bacteria in the organs and succumbed to infection much earlier than C57BL/6 animals. Absence of MyD88 resulted in diminished production of inflammatory cytokines such as interleukin-12, interferon-γ, and tumor necrosis factor-α as well as chemoattractants such as monocyte chemotactic protein-1 (MCP-1) and Keratinocyte-derived chemokine (KC), and hampered recruitment of effector cells involved in bacterial clearance (macrophages and neutrophils) to the infection site. Furthermore, MyD88−/− but not C57BL/6 mice exhibited a massive infiltration of eosinophils in infected organs, which can be explained by an impaired production of the regulatory chemokines, gamma interferon-induced monokine (MIG/CXCL9) and interferon-induced protein 10 (IP-10/CXCL10), which can inhibit transmigration of eosinophils. Our results indicate that MyD88 signaling targets effector cells to the site of streptococcal infection and prevents extravasation of cells that can induce tissue damage. Therefore, MyD88 signaling may be important for shaping the quality of the inflammatory response elicited during infection to ensure optimal effector functions. PMID:20019195

  2. Transport of. cap alpha. -aminoisobutyric acid by Streptococcus pyogenes and its derived L-form

    SciTech Connect

    Reizer, J.; Panos, C.

    1982-01-01

    We studied the uptake of ..cap alpha..-aminoisobutyric acid (AIB) in Streptococcus pyogenes and its physiologically isotonic L-form. S. pyogenes cells starved for glucose or treated with carbonyl cyanide-m-chlorophenyl hydrazone accumulated limited amounts of AIB. A high apparent K/sub m/ value characterized the glucose-independent transport of AIB. The rate and extent of AIB accumulation significantly increased in the presence of glucose. Two saturable transport components with distinct apparent K/sub m/values characterized glycolysis-coupled transport of AIB. A biphasic Lineweaver-Burk plot was also obtained for L-alanine transport by glycolyzing S. pyogenes cells. AIB seems to share a common transport system(s) with glycine, L- and D-anine, L-serine, and L-valine. This was shown by the competitive exchange efflux of accumulated AIB. About 30% of the AIB uptake was not inhibited by a saturating amount of L-valine, indicating the existence of more than one system for AIB transport, p-Chloromercuribenzoate markedly inhibited the accumulation of AIB by both glycolyzing and glucose-starved cells. In contrast, carbonyl cyanide-m-chlorophenyl hydrazone affected only metabolism-dependent uptake of AIB, which was also sensitive to dinitrophenol, N-ethylmaleimide, iodoacetate, fluoride (NaF), arsenate, and N,N'-dicyclohexylcarbodiimide. These results are interpreted according to the chemiosmotic theory of Mitchell, whereby a proton motive force constitutes the driving force for AIB accumulation. AIB was not accumulated by the L-form. However, a temporary accumulation of AIB by a counterflow mechanism and a saturable system with a low apparent affinity were demonstrated for AIB transport by this organism. We suggest that a deficiency in the coupling of energy to AIB transport is responsible for the apparent lack of active AIB accumulation by the L-form.

  3. Loss of erythromycin resistance genes from strains of Streptococcus pyogenes that have developed resistance to levofloxacin.

    PubMed

    Billal, Dewan Sakhawat; Hotomi, Muneki; Yan, Steve S; Fedorko, Daniel P; Shimada, Jun; Fujihara, Keiji; Yamanaka, Noboru

    2009-06-01

    In the past 2 to 3 decades, erythromycin resistance in Streptococcus pyogenes has been decreasing, whereas fluoroquinolone resistance (or reduction in its susceptibility) has been reported often. Although a shift of M-type prevalence and decreased pressure from macrolides have been suggested for the decrease in erythromycin resistance, we hypothesized that this might also be a result of increased antimicrobial pressure from fluoroquinolone use. Levofloxacin resistance for 4 erythromycin-resistant parent strains was induced in vitro. Their mutants became highly resistant to the fluoroquinolones but lost their erythromycin resistance trait. Erythromycin resistance was fully restored by transconjugation with respective parent strains with either mefA- or ermTR-mediated mechanisms.

  4. Scrum kidney: epidemic pyoderma caused by a nephritogenic Streptococcus pyogenes in a rugby team.

    PubMed

    Ludlam, H; Cookson, B

    1986-08-09

    In December, 1984, an outbreak of pyoderma affected five scrum players in the St Thomas' Hospital rugby team. The causative organism, Streptococcus pyogenes, was acquired during a match against a team experiencing an outbreak of impetigo, and was transmitted to two front row players of another team a week later, and to two girlfriends of affected St Thomas' players a month later. The strain was M-type 49, tetracycline-resistant, and virulent. It caused salpingitis in a girlfriend and acute glomerulonephritis in one rugby player. No case of subclinical glomerulonephritis was detected in eight patients with pyoderma. Screening of the St Thomas' Hospital team revealed four further cases of non-streptococcal skin infection, with evidence for contemporaneous spread of Staphylococcus aureus. Teams should not field players with sepsis, and it may be advisable to apply a skin antiseptic to traumatised skin after the match.

  5. An intrinsic pattern of reduced susceptibility to fluoroquinolones in pediatric isolates of Streptococcus pyogenes.

    PubMed

    Yan, S Steve; Schreckenberger, Paul C; Zheng, Xiaotian; Nelson, Nancy A; Harrington, Susan M; Tjhio, Joyce; Fedorko, Daniel P

    2008-10-01

    A total of 116 clinical isolates collected in 2003 from a tertiary pediatric hospital and a primary pediatric department in Chicago, IL, were screened for reduced susceptibility to selected fluoroquinolones by disc diffusion. Correlation between reduced susceptibility and point mutations in the quinolone resistance-determining region of parC and gyrA genes was evaluated, and point mutations were compared with other reports of isolates derived from adult or mixed patient populations. Nine percent of isolates had reduced susceptibility to 1 or more of these fluoroquinolones by Etest: ciprofloxacin, levofloxacin, and moxifloxacin. A single point mutation (Ser-79) in parC seemed responsible for the reduced susceptibility. Resistant Streptococcus pyogenes isolates were compared using M/emm type, repetitive sequence-based PCR (rep-PCR), and pulsed-field gel electrophoresis (PFGE). Rep-PCR provided no more separation of strains than M/emm typing, and PFGE results with SgrAI were more discriminatory than with SmaI. The majority of these isolates were M/emm type 6. PFGE analysis using SgrAI demonstrated 2 different resistant strains among the M/emm type 6 isolates. The findings suggest that a population of S. pyogenes with an intrinsic reduced susceptibility to fluoroquinolones exists in pediatric clinical isolates. Monitoring of amino acid changes in both parC and gyrA will assist in the prediction of emergence of high-level fluoroquinolone resistance.

  6. Structural and functional characterization of Streptococcus pyogenes Cas2 protein under different pH conditions.

    PubMed

    Ka, Donghyun; Kim, Dayoun; Baek, Gyeongyun; Bae, Euiyoung

    2014-08-15

    Clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins constitute an RNA-guided microbial defense system against invading foreign genetic materials. Cas2 is one of the core Cas proteins found universally in all the subtypes of CRISPR-Cas systems and is required for incorporating new spacers into CRISPR loci. Cas2 homologues from different CRISPR-Cas subtypes were characterized previously as metal-dependent nucleases with different substrate preferences, and it was proposed that a pH-dependent conformational change mediates metal binding and catalysis. Here, we report the crystal structures of Streptococcus pyogenes Cas2 at three different pHs (5.6, 6.5, and 7.5), as well as the results of its nuclease activity assay against double-stranded DNAs at varying pHs (6.0-9.0). Although S. pyogenes Cas2 exhibited strongly pH-dependent catalytic activity, there was no significant conformational difference among the three crystal structures. However, structural comparisons with other Cas2 homologues revealed structural variability and the flexible nature of its putative hinge regions, supporting the hypothesis that conformational switching is important for catalysis. Taken together, our results confirm that Cas2 proteins have pH-dependent nuclease activity against double-stranded DNAs, and provide indirect structural evidence for their conformational changes. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. A vaccine against Streptococcus pyogenes: the potential to prevent rheumatic fever and rheumatic heart disease.

    PubMed

    Guilherme, Luiza; Ferreira, Frederico Moraes; Köhler, Karen Francine; Postol, Edilberto; Kalil, Jorge

    2013-02-01

    Streptococcus pyogenes causes severe, invasive infections such as the sequelae associated with acute rheumatic fever, rheumatic heart disease, acute glomerulonephritis, uncomplicated pharyngitis, and pyoderma. Efforts to produce a vaccine against S. pyogenes began several decades ago, and different models have been proposed. We have developed a vaccine candidate peptide, StreptInCor, comprising 55 amino acid residues of the C-terminal portion of the M protein and encompassing both the T- and B-cell protective epitopes. The present article summarizes data from the previous 5 years during which we tested the immunogenicity and safety of StreptInCor in different animal models. We showed that StreptInCor overlapping peptides induced cellular and humoral immune responses of individuals bearing different HLA class II molecules. These results are consistent with peptides that have a universal vaccine epitope. The tridimensional molecular structure of StreptInCor was elucidated by nuclear magnetic resonance spectroscopy, which showed that its structure is composed of two microdomains linked by an 18-residue α-helix. Additionally, we comprehensively evaluated the structural stability of the StreptInCor peptide in different physicochemical conditions using circular dichroism. Additional experiments were performed with inbred, outbred, and HLA class II transgenic mice. Analysis of several organs of these mice showed neither deleterious nor autoimmune reactions even after a long period of vaccination, indicating that the StreptInCor candidate peptide could be considered as an immunogenic and safe vaccine.

  8. A study on erm(B)-mediated MLS resistance in Streptococcus pyogenes clinical isolates.

    PubMed

    Rombini, Silvia; Petrelli, Dezemona; Bolli, Elisabetta; Tran, Chi Nhan; Falconi, Maurizio; Di Luca, Maria Chiara; Prenna, Manuela; Ripa, Sandro; Vitali, Luca Agostino

    2011-07-01

    The constitutive or inducible macrolide-lincosamide-streptogramin (MLS) phenotype of 30 erm(B)-positive Streptococcus pyogenes isolates was determined by different methods and under various growth conditions and correlated to the sequence of the 5'-untranslated regions of erm(B). The MLS phenotype of one-third of the isolates could not be classified. In liquid medium, some of these isolates responded to induction only during the logarithmic phase of growth, while others expressed clindamycin resistance even under noninducing conditions. By increasing the growth rate, we observed a shift from a constitutive towards an inducible pattern of resistance. All data were confirmed by analysis of the 23S rRNA methylation level. The erm(B)-5'-untranslated region was 99% similar in sequence. In erm(B)-positive S. pyogenes, the MLS phenotype is strongly influenced by culture conditions and control of its expression does not depend exclusively on the sequence of the erm(B)-5'-untranslated region. Copyright © 2011 Elsevier Inc. All rights reserved.

  9. Relationships between emm and multilocus sequence types within a global collection of Streptococcus pyogenes.

    PubMed

    Bessen, Debra E; McGregor, Karen F; Whatmore, Adrian M

    2008-04-11

    The M type-specific surface protein antigens encoded by the 5' end of emm genes are targets of protective host immunity and attractive vaccine candidates against infection by Streptococcus pyogenes, a global human pathogen. A history of genetic change in emm was evaluated for a worldwide collection of > 500 S. pyogenes isolates that were defined for genetic background by multilocus sequence typing of housekeeping genes. Organisms were categorized by genotypes that roughly correspond to throat specialists, skin specialists, and generalists often recovered from infections at either tissue site. Recovery of distant clones sharing the same emm type was approximately 4-fold higher for skin specialists and generalists, as compared to throat specialists. Importantly, emm type was often a poor marker for clone. Recovery of clones that underwent recombinational replacement with a new emm type was most evident for the throat and skin specialists. The average ratio of nonsynonymous substitutions per nonsynonymous site (Ka) and synonymous substitutions per synonymous site (Ks) was 4.9, 1.5 and 1.3 for emm types of the throat specialist, skin specialist and generalist groups, respectively. Data indicate that the relationships between emm type and genetic background differ among the three host tissue-related groups, and that the selection pressures acting on emm appear to be strongest for the throat specialists. Since positive selection is likely due in part to a protective host immune response, the findings may have important implications for vaccine design and vaccination strategies.

  10. Susceptibility and emm type of Streptococcus pyogenes isolated from children with severe infection.

    PubMed

    Sakata, Hiroshi

    2013-12-01

    Minimal inhibitory concentrations (MICs) and minimal bactericidal concentrations (MBCs) of various antimicrobial agents were measured against 12 strains of Streptococcus pyogenes isolated from children with invasive infections between 2003 and 2012. The patients ranged in age from 1 day to 15 years, with patients younger than 5 years, including three neonates, accounting for a half of the patients. The disease was sepsis in four patients, skin and soft tissue infection in three patients, retropharyngeal abscess in two patients, pneumonia plus sepsis in one patient, empyema in one patient, and pyogenic arthritis in one patient. One patient with sepsis died, while cure without sequelae was achieved in all the remaining patients. When classified by type, emm1 (six strains) was the most prevalent type, followed by emm12 (two strains). The MIC90/MBC90 values were 0.015/0.015 μg/mL for penicillin G, 0.03/0.03 μg/mL for ampicillin, 0.015/0.03 μg/mL for cefotaxime, 0.03/0.03 μg/mL for ceftriaxone, 0.008/0.008 μg/mL for panipenem, 0.008/0.008 μg/mL for meropenem, and ≤0.004/≤0.004 μg/mL for doripenem, indicating the superior antimicrobial activities of carbapenem.

  11. Sequence variability is correlated with weak immunogenicity in Streptococcus pyogenes M protein

    PubMed Central

    Lannergård, Jonas; Kristensen, Bodil M; Gustafsson, Mattias C U; Persson, Jenny J; Norrby-Teglund, Anna; Stålhammar-Carlemalm, Margaretha; Lindahl, Gunnar

    2015-01-01

    The M protein of Streptococcus pyogenes, a major bacterial virulence factor, has an amino-terminal hypervariable region (HVR) that is a target for type-specific protective antibodies. Intriguingly, the HVR elicits a weak antibody response, indicating that it escapes host immunity by two mechanisms, sequence variability and weak immunogenicity. However, the properties influencing the immunogenicity of regions in an M protein remain poorly understood. Here, we studied the antibody response to different regions of the classical M1 and M5 proteins, in which not only the HVR but also the adjacent fibrinogen-binding B repeat region exhibits extensive sequence divergence. Analysis of antisera from S. pyogenes-infected patients, infected mice, and immunized mice showed that both the HVR and the B repeat region elicited weak antibody responses, while the conserved carboxy-terminal part was immunodominant. Thus, we identified a correlation between sequence variability and weak immunogenicity for M protein regions. A potential explanation for the weak immunogenicity was provided by the demonstration that protease digestion selectively eliminated the HVR-B part from whole M protein-expressing bacteria. These data support a coherent model, in which the entire variable HVR-B part evades antibody attack, not only by sequence variability but also by weak immunogenicity resulting from protease attack. PMID:26175306

  12. SpeB of Streptococcus pyogenes Differentially Modulates Antibacterial and Receptor Activating Properties of Human Chemokines

    PubMed Central

    Egesten, Arne; Olin, Anders I.; Linge, Helena M.; Yadav, Manisha; Mörgelin, Matthias; Karlsson, Anna; Collin, Mattias

    2009-01-01

    Background CXC chemokines are induced by inflammatory stimuli in epithelial cells and some, like MIG/CXCL9, IP–10/CXCL10 and I–TAC/CXCL11, are antibacterial for Streptococcus pyogenes. Methodology/Principal Findings SpeB from S. pyogenes degrades a wide range of chemokines (i.e. IP10/CXCL10, I-TAC/CXCL11, PF4/CXCL4, GROα/CXCL1, GROβ/CXCL2, GROγ/CXCL3, ENA78/CXCL5, GCP-2/CXCL6, NAP-2/CXCL7, SDF-1/CXCL12, BCA-1/CXCL13, BRAK/CXCL14, SRPSOX/CXCL16, MIP-3α/CCL20, Lymphotactin/XCL1, and Fractalkine/CX3CL1), has no activity on IL-8/CXCL8 and RANTES/CCL5, partly degrades SRPSOX/CXCL16 and MIP-3α/CCL20, and releases a 6 kDa CXCL9 fragment. CXCL10 and CXCL11 loose receptor activating and antibacterial activities, while the CXCL9 fragment does not activate the receptor CXCR3 but retains its antibacterial activity. Conclusions/Significance SpeB destroys most of the signaling and antibacterial properties of chemokines expressed by an inflamed epithelium. The exception is CXCL9 that preserves its antibacterial activity after hydrolysis, emphasizing its role as a major antimicrobial on inflamed epithelium. PMID:19274094

  13. [Susceptibility of Streptococcus pyogenes to macrolides and quinolones in Guadalajara, Spain].

    PubMed

    Rodríguez-Zurita, M E; Solís del Baño, S; Robres Guillén, P; González Praetorius, A; Gimeno Fernández, C; Pérez Pomata, M T; Bisquert Santiago, J

    2003-03-01

    We carried out a retrospective study of the susceptibility of 104 Streptococcus pyogenes strains, which were isolated in 2000 and 2001 from clinical samples of different origins, to penicillin, erythromycin, clindamycin, ofloxacin and levofloxacin. The susceptibility testing was performed using the agar difusion method according to the guidelines of the NCCLS. All of the isolates showed susceptibility to penicillin and clindamycin. However, we detected 11 strains that were resistant to erythromycin (10.6%) and 4 strains resistant to ofloxacin (3.8%). We studied the resistance phenotypes of macrolides and lincosamides using erythromycin and clindamycin discs. Nine of the eleven strains that were resistant to erythromycin showed an M phenotype, while the remaining two showed inducible resistance to clindamycin, thus suggesting an MLS(B) inducible phenotype. No strains with constitutive resistance to erythromycin or clindamycin (MLS(B) constitutive phenotype) were identified. While penicillin is still uniformly active against S. pyogenes, in Guadalajara, there are 10.6% strains that are resistant to 14- and 15-atoms macrolides.

  14. emm Gene distribution among erythromycin-resistant and -susceptible Italian isolates of Streptococcus pyogenes.

    PubMed

    Zampaloni, Claudia; Cappelletti, Paola; Prenna, Manuela; Vitali, Luca Agostino; Ripa, Sandro

    2003-03-01

    The phenotypes and genetic determinants for macrolide resistance were determined for 167 erythromycin-resistant Streptococcus pyogenes strains. A cMLS phenotype was shown in 18% of the erythromycin-resistant strains, while inducible resistance was apparent in 31% and the M phenotype was apparent in 50%. The emm gene type of this set of resistant isolates and that of 48 erythromycin-sensitive isolates were determined. emm2 and emm48 were recorded only in the resistant strains of the M phenotype, while approximately all of the strains harboring the emm22 gene had the cMLS phenotype. More than 80% of the emm89-positive strains had the iMLS phenotype, and the same portion of emm4 strains presented the M phenotype. emm3 is recorded only among sensitive strains. The distribution of frequencies of the genetic determinant for the virulence factor M protein was significantly different both among organisms of different types of resistance and between resistant and sensitive populations of S. pyogenes under study.

  15. emm Gene Distribution among Erythromycin-Resistant and -Susceptible Italian Isolates of Streptococcus pyogenes

    PubMed Central

    Zampaloni, Claudia; Cappelletti, Paola; Prenna, Manuela; Vitali, Luca Agostino; Ripa, Sandro

    2003-01-01

    The phenotypes and genetic determinants for macrolide resistance were determined for 167 erythromycin-resistant Streptococcus pyogenes strains. A cMLS phenotype was shown in 18% of the erythromycin-resistant strains, while inducible resistance was apparent in 31% and the M phenotype was apparent in 50%. The emm gene type of this set of resistant isolates and that of 48 erythromycin-sensitive isolates were determined. emm2 and emm48 were recorded only in the resistant strains of the M phenotype, while approximately all of the strains harboring the emm22 gene had the cMLS phenotype. More than 80% of the emm89-positive strains had the iMLS phenotype, and the same portion of emm4 strains presented the M phenotype. emm3 is recorded only among sensitive strains. The distribution of frequencies of the genetic determinant for the virulence factor M protein was significantly different both among organisms of different types of resistance and between resistant and sensitive populations of S. pyogenes under study. PMID:12624074

  16. Macrolide and Tetracycline Resistance and emm Type Distribution of Streptococcus pyogenes Isolates Recovered from Turkish Patients

    PubMed Central

    Sayan, Murat; Tamer, Gulden Sonmez

    2010-01-01

    The aims of this study were to determine the susceptibilities to macrolide and tetracycline antibiotics and emm type distribution of Streptococcus pyogenes strains isolated in the Kocaeli University Hospital, Turkey. A total of 127 S. pyogenes clinical isolates were tested. Eleven (9%) isolates were resistant to erythromycin, and 23 (18%) isolates were resistant to tetracycline. Ten of the erythromycin-resistant isolates were also resistant to tetracycline. By the triple-disk test, all erythromycin-resistant isolates showed the inducible macrolide-lincosamide-streptogramin-C phenotype and harbored erm(TR) gene. tet(O) was the most common tetracycline resistance gene. Among erythromycin-tetracycline coresistant isolates, seven harbored the tet(O) gene. emm 4, emm 1, emm 2,114, and emm 89 were the most common emm types. These isolates were more susceptible to erythromycin. There was considerable emm type heterogeneity in macrolide or tetracycline resistant isolates. According to our knowledge, this is the first study in which emm type distribution is investigated in Turkey. More comprehensive studies are needed to obtain true information about the epidemiology of macrolide and tetracycline resistance and emm type distribution in Turkey. PMID:20624096

  17. Inactivated and live, attenuated influenza vaccines protect mice against influenza:Streptococcus pyogenes super-infections

    PubMed Central

    Chaussee, Michael S.; Sandbulte, Heather R.; Schuneman, Margaret J.; DePaula, Frank P.; Addengast, Leslie A.; Schlenker, Evelyn H.; Huber, Victor C.

    2011-01-01

    Mortality associated with influenza virus super-infections is frequently due to secondary bacterial complications. To date, super-infections with Streptococcus pyogenes have been studied less extensively than those associated with S. pneumoniae. This is significant because a vaccine for S. pyogenes is not clinically available, leaving vaccination against influenza virus as our only means for preventing these super-infections. In this study, we directly compared immunity induced by two types of influenza vaccine, either inactivated influenza virus (IIV) or live, attenuated influenza virus (LAIV), for the ability to prevent super-infections. Our data demonstrate that both IIV and LAIV vaccines induce similar levels of serum antibodies, and that LAIV alone induces IgA expression at mucosal surfaces. Upon super-infection, both vaccines have the ability to limit the induction of pro-inflammatory cytokines within the lung, including IFN-γ which has been shown to contribute to mortality in previous models of super-infection. Limiting expression of these pro-inflammatory cytokines within the lungs subsequently limits recruitment of macrophages and neutrophils to pulmonary surfaces, and ultimately protects both IIV- and LAIV-vaccinated mice from mortality. Despite their overall survival, both IIV- and LAIV-vaccinated mice demonstrated levels of bacteria within the lung tissue to levels that are similar to those seen in unvaccinated mice. Thus, influenza virus:bacteria super-infections can be limited by vaccine-induced immunity against influenza virus, but the ability to prevent morbidity is not complete. PMID:21440037

  18. [Detection of early gastric cancer facilitated by surveillance for a pyogenic liver abscess caused by Streptococcus intermedius].

    PubMed

    Shigefuku, Ryuta; Matsunaga, Kotaro; Tamura, Tomohiro; Ozawa, Shun-Ichiro; Matsuo, Yasumasa; Takahashi, Hideaki; Matsumoto, Nobuyuki; Okuse, Chiaki; Suzuki, Michihiro; Itoh, Fumio

    2016-01-01

    We report a case of early gastric cancer that was detected during surveillance of a pyogenic liver abscess caused by Streptococcus intermedius, an oral microbiota. Treatment with proton pump inhibitors can result in the alteration of gastric bacterial flora by altering intragastric acidity. This can place immunocompromised patients, such as those with diabetes mellitus and the elderly, at an increased risk for disease of the upper gastrointestinal tract to be a route of bacterial transmission. In this case, the patient developed a pyogenic liver abscess.

  19. Draft Genome Sequence of Streptococcus pyogenes Strain 06BA18369, a Human Pathogen Associated with Skin and Soft Tissue Infections in Northern Canada.

    PubMed

    McDonald, Ryan R; Golding, George R; Irvine, James; Graham, Morag R; Tyler, Shaun; Mulvey, Michael R; Levett, Paul N

    2013-06-27

    We report the draft sequence of Streptococcus pyogenes 06BA18369 (emm type 41.2, sequence type 579 [ST579]), isolated from a skin and soft tissue infection (SSTI) mixed with Staphylococcus aureus. This genome provides insight into the genetic composition of S. pyogenes strains associated with mixed SSTIs.

  20. An improved SELEX technique for selection of DNA aptamers binding to M-type 11 of Streptococcus pyogenes.

    PubMed

    Hamula, Camille L A; Peng, Hanyong; Wang, Zhixin; Tyrrell, Gregory J; Li, Xing-Fang; Le, X Chris

    2016-03-15

    Streptococcus pyogenes is a clinically important pathogen consisting of various serotypes determined by different M proteins expressed on the cell surface. The M type is therefore a useful marker to monitor the spread of invasive S. pyogenes in a population. Serotyping and nucleic acid amplification/sequencing methods for the identification of M types are laborious, inconsistent, and usually confined to reference laboratories. The primary objective of this work is to develop a technique that enables generation of aptamers binding to specific M-types of S. pyogenes. We describe here an in vitro technique that directly used live bacterial cells and the Systematic Evolution of Ligands by Exponential Enrichment (SELEX) strategy. Live S. pyogenes cells were incubated with DNA libraries consisting of 40-nucleotides randomized sequences. Those sequences that bound to the cells were separated, amplified using polymerase chain reaction (PCR), purified using gel electrophoresis, and served as the input DNA pool for the next round of SELEX selection. A specially designed forward primer containing extended polyA20/5Sp9 facilitated gel electrophoresis purification of ssDNA after PCR amplification. A counter-selection step using non-target cells was introduced to improve selectivity. DNA libraries of different starting sequence diversity (10(16) and 10(14)) were compared. Aptamer pools from each round of selection were tested for their binding to the target and non-target cells using flow cytometry. Selected aptamer pools were then cloned and sequenced. Individual aptamer sequences were screened on the basis of their binding to the 10 M-types that were used as targets. Aptamer pools obtained from SELEX rounds 5-8 showed high affinity to the target S. pyogenes cells. Tests against non-target Streptococcus bovis, Streptococcus pneumoniae, and Enterococcus species demonstrated selectivity of these aptamers for binding to S. pyogenes. Several aptamer sequences were found to bind

  1. Functional and Structural Properties of a Novel Protein and Virulence Factor (Protein sHIP) in Streptococcus pyogenes *

    PubMed Central

    Wisniewska, Magdalena; Happonen, Lotta; Kahn, Fredrik; Varjosalo, Markku; Malmström, Lars; Rosenberger, George; Karlsson, Christofer; Cazzamali, Giuseppe; Pozdnyakova, Irina; Frick, Inga-Maria; Björck, Lars; Streicher, Werner; Malmström, Johan; Wikström, Mats

    2014-01-01

    Streptococcus pyogenes is a significant bacterial pathogen in the human population. The importance of virulence factors for the survival and colonization of S. pyogenes is well established, and many of these factors are exposed to the extracellular environment, enabling bacterial interactions with the host. In the present study, we quantitatively analyzed and compared S. pyogenes proteins in the growth medium of a strain that is virulent to mice with a non-virulent strain. Particularly, one of these proteins was present at significantly higher levels in stationary growth medium from the virulent strain. We determined the three-dimensional structure of the protein that showed a unique tetrameric organization composed of four helix-loop-helix motifs. Affinity pull-down mass spectrometry analysis in human plasma demonstrated that the protein interacts with histidine-rich glycoprotein (HRG), and the name sHIP (streptococcal histidine-rich glycoprotein-interacting protein) is therefore proposed. HRG has antibacterial activity, and when challenged by HRG, sHIP was found to rescue S. pyogenes bacteria. This and the finding that patients with invasive S. pyogenes infection respond with antibody production against sHIP suggest a role for the protein in S. pyogenes pathogenesis. PMID:24825900

  2. Role of serine/threonine phosphatase (SP-STP) in Streptococcus pyogenes physiology and virulence.

    PubMed

    Agarwal, Shivani; Agarwal, Shivangi; Pancholi, Preeti; Pancholi, Vijay

    2011-12-02

    Reversible phosphorylation is the key mechanism regulating several cellular events in prokaryotes and eukaryotes. In prokaryotes, signal transduction is perceived to occur primarily via the two-component signaling system involving histidine kinases and cognate response regulators. Although an alternative regulatory pathway controlled by the eukaryote-type serine/threonine kinase (Streptococcus pyogenes serine/threonine kinase; SP-STK) has been shown to modulate bacterial growth, division, adherence, invasion, and virulence in group A Streptococcus (GAS; S. pyogenes), the precise role of the co-transcribing serine/threonine phosphatase (SP-STP) has remained enigmatic. In this context, this is the first report describing the construction and characterization of non-polar SP-STP mutants in two different strains of Type M1 GAS. The STP knock-out mutants displayed increased bacterial chain lengths in conjunction with thickened cell walls, significantly reduced capsule and hemolysin production, and restoration of the phenotypes postcomplementation. The present study also reveals important contribution of cognately regulated-reversible phosphorylation by SP-STK/SP-STP on two major response regulators of two-component systems, WalRK and CovRS. We also demonstrate a distinct role of SP-STP in terms of expression of surface proteins and SpeB in a strain-specific manner. Further, the attenuation of virulence in the absence of STP and its restoration only in the complemented strains that were generated by the use of a low copy plasmid and not by a high copy one emphasize not only the essential role of STP in virulence but also highlight the tightly regulated SP-STP/SP-STK-mediated cognate functions. SP-STP thus is an important regulator of GAS virulence and plays a critical role in GAS pathogenesis.

  3. Role of Serine/Threonine Phosphatase (SP-STP) in Streptococcus pyogenes Physiology and Virulence*

    PubMed Central

    Agarwal, Shivani; Agarwal, Shivangi; Pancholi, Preeti; Pancholi, Vijay

    2011-01-01

    Reversible phosphorylation is the key mechanism regulating several cellular events in prokaryotes and eukaryotes. In prokaryotes, signal transduction is perceived to occur primarily via the two-component signaling system involving histidine kinases and cognate response regulators. Although an alternative regulatory pathway controlled by the eukaryote-type serine/threonine kinase (Streptococcus pyogenes serine/threonine kinase; SP-STK) has been shown to modulate bacterial growth, division, adherence, invasion, and virulence in group A Streptococcus (GAS; S. pyogenes), the precise role of the co-transcribing serine/threonine phosphatase (SP-STP) has remained enigmatic. In this context, this is the first report describing the construction and characterization of non-polar SP-STP mutants in two different strains of Type M1 GAS. The STP knock-out mutants displayed increased bacterial chain lengths in conjunction with thickened cell walls, significantly reduced capsule and hemolysin production, and restoration of the phenotypes postcomplementation. The present study also reveals important contribution of cognately regulated-reversible phosphorylation by SP-STK/SP-STP on two major response regulators of two-component systems, WalRK and CovRS. We also demonstrate a distinct role of SP-STP in terms of expression of surface proteins and SpeB in a strain-specific manner. Further, the attenuation of virulence in the absence of STP and its restoration only in the complemented strains that were generated by the use of a low copy plasmid and not by a high copy one emphasize not only the essential role of STP in virulence but also highlight the tightly regulated SP-STP/SP-STK-mediated cognate functions. SP-STP thus is an important regulator of GAS virulence and plays a critical role in GAS pathogenesis. PMID:21917918

  4. Inactivated and live, attenuated influenza vaccines protect mice against influenza: Streptococcus pyogenes super-infections.

    PubMed

    Chaussee, Michael S; Sandbulte, Heather R; Schuneman, Margaret J; Depaula, Frank P; Addengast, Leslie A; Schlenker, Evelyn H; Huber, Victor C

    2011-05-12

    Mortality associated with influenza virus super-infections is frequently due to secondary bacterial complications. To date, super-infections with Streptococcus pyogenes have been studied less extensively than those associated with Streptococcus pneumoniae. This is significant because a vaccine for S. pyogenes is not clinically available, leaving vaccination against influenza virus as our only means for preventing these super-infections. In this study, we directly compared immunity induced by two types of influenza vaccine, either inactivated influenza virus (IIV) or live, attenuated influenza virus (LAIV), for the ability to prevent super-infections. Our data demonstrate that both IIV and LAIV vaccines induce similar levels of serum antibodies, and that LAIV alone induces IgA expression at mucosal surfaces. Upon super-infection, both vaccines have the ability to limit the induction of pro-inflammatory cytokines within the lung, including IFN-γ which has been shown to contribute to mortality in previous models of super-infection. Limiting expression of these pro-inflammatory cytokines within the lungs subsequently limits recruitment of macrophages and neutrophils to pulmonary surfaces, and ultimately protects both IIV- and LAIV-vaccinated mice from mortality. Despite their overall survival, both IIV- and LAIV-vaccinated mice demonstrated levels of bacteria within the lung tissue that are similar to those seen in unvaccinated mice. Thus, influenza virus:bacteria super-infections can be limited by vaccine-induced immunity against influenza virus, but the ability to prevent morbidity is not complete. Copyright © 2011 Elsevier Ltd. All rights reserved.

  5. Primary peritonitis by Streptococcus pyogenes. A condition as rare as it is aggressive.

    PubMed

    Abellán Morcillo, Israel; González, Antonio; Selva Cabañero, Pilar; Bernabé, Antonio

    2016-04-01

    We report the case of a 60-year-old female patient who presented to the emergency room for abdominal pain standing with impaired general status, fever of up to 38.7ºC, and somnolence. Upon arrival the patient had a heart rate of 115 bpm, hypotension (80/40 mmHg),acute respiratory distress, and both hepatic and renal failure. During her examination the patient was drowsy and had a diffusely tender abdomen with peritoneal irritation signs. Blood tests revealed 22,000 WBCs (82%N), CRP 32.4 mg/dL, total bilirubin 3.2 mg/dL, GOT 300 U/L, GPT 160 U/L, LDH 200 U/L, AP 310 U/L, 91,000 platelets, creatinine2.3 mg/dL, and PA 64%. An abdominal CT scan was performed, which revealed a minimal amount of free intraperitoneal fluid with no other findings. Given the patient's poor status an exploratory laparoscopy was carried out, which found a moderate amount of diffuse purulent exudate, particularly in interloop and lesser pelvis areas, with no additional findings. Following surgery she was transferred to the intensive care unit on wide spectrum antibiotics .Peritoneal exudate cultures from the surgical procedure revealed Streptococcus pyogenes. The patient had a favorable outcome being subsequently discharged from hospital at day 10 after the procedure. S. pyogenesis a beta hemolytic streptococcus well known as a cause of pharyngotonsillar, skin and soft tissues infection. Primary peritonitis by S.pyogenesis a rare condition with only a few isolated cases reported. PP cases by S.pyogenes predominantly involve previously healthy young women. PP diagnosis is usually retrospective, when other causes have been ruled out by surgery and culture is positive post hoc. An appropriate differential diagnosis from conditions such as gram-negative shock, staphylococcal toxic shock, meningococcal disease, viral infection, etc., is crucial. Abdominal CT may be helpful but a variable amount of free intraperitoneal fluid is usually the only finding. The surgical approach is usually laparoscopy

  6. Telithromycin and Quinupristin-Dalfopristin Resistance in Clinical Isolates of Streptococcus pyogenes: SMART Program 2001 Data

    PubMed Central

    Hsueh, Po-Ren; Teng, Lee-Jene; Lee, Chun-Ming; Huang, Wen-Kuei; Wu, Tsu-Lan; Wan, Jen-Hsien; Yang, Dine; Shyr, Jainn-Ming; Chuang, Yin-Ching; Yan, Jing-Jou; Lu, Jang-Jih; Wu, Jiunn-Jong; Ko, Wen-Chien; Chang, Feng-Yee; Yang, Yi-Chueh; Lau, Yeu-Jun; Liu, Yung-Ching; Leu, Hsieh-Shong; Liu, Cheng-Yi; Luh, Kwen-Tay

    2003-01-01

    This study evaluated the current status of antimicrobial resistance in clinical isolates of Streptococcus pyogenes in Taiwan as part of the SMART (Surveillance from Multicenter Antimicrobial Resistance in Taiwan) program. In 2001, 419 different isolates of S. pyogenes, including 275 from respiratory secretions, 87 from wound pus, and 31 from blood, were collected from nine hospitals in different parts of Taiwan. MICs of 23 antimicrobial agents were determined at a central location by the agar dilution method. All of the isolates were susceptible to penicillin (MIC at which 90% of the isolates were inhibited [MIC90], ≤0.03 μg/ml), cefotaxime (MIC90, ≤0.03 μg/ml), cefepime (MIC90, 0.06 μg/ml), meropenem (MIC90, ≤0.03 μg/ml), moxifloxacin (MIC90, 0.25 μg/ml), vancomycin (MIC90, 0.5 μg/ml), and linezolid (MIC90, 1 μg/ml). Overall, 78% of isolates were not susceptible to erythromycin (54% were intermediate, and 24% were resistant), and 5% were not susceptible to clindamycin. Of the 101 erythromycin-resistant isolates, 80.2% exhibited the M phenotype (mefA gene positive), 18.9% exhibited the cMLS (constitutive resistance to macrolides-lincosamides-streptogramin B [MLS]) phenotype (ermB gene positive), and 1% exhibited the iMLS (inducible resistance to MLS) phenotype (ermB gene positive). Fluoroquinolones (sitafloxacin > moxifloxacin > ciprofloxacin = levofloxacin = gatifloxacin > gemifloxacin) demonstrated potent activity against nearly all of the isolates of S. pyogenes tested. Thirty-two isolates (8%) were not susceptible to quinupristin-dalfopristin. Seventeen percent of isolates had telithromycin MICs of ≥1 μg/ml, and all of these isolates exhibited erythromycin MICs of ≥32 μg/ml. The high prevalence of resistance to telithromycin (which is not available in Taiwan) limits its potential use in the treatment of S. pyogenes infections, particularly in areas with high rates of macrolide resistance. PMID:12821461

  7. Streptokinase variants from Streptococcus pyogenes isolates display altered plasminogen activation characteristics - implications for pathogenesis.

    PubMed

    Cook, Simon M; Skora, Amanda; Gillen, Christine M; Walker, Mark J; McArthur, Jason D

    2012-12-01

    Streptococcus pyogenes (group A streptococcus, GAS) secretes streptokinase, a potent plasminogen activating protein. Among GAS isolates, streptokinase gene sequences (ska) are polymorphic and can be grouped into two distinct sequence clusters (termed cluster type-1 and cluster type-2) with cluster type-2 being further divided into sub-clusters type-2a and type-2b. In this study, far-UV circular dichroism spectroscopy indicated that purified streptokinase variants of each type displayed similar secondary structure. Type-2b streptokinase variants could not generate an active site in Glu-plasminogen through non-proteolytic mechanisms while all other variants had this capability. Furthermore, when compared with other streptokinase variants, type-2b variants displayed a 29- to 35-fold reduction in affinity for Glu-plasminogen. All SK variants could activate Glu-plasminogen when an activator complex was preformed with plasmin; however, type-2b and type-1 complexes were inhibited by α(2) -antiplasmin. Exchanging ska(type-2a) in the M1T1 GAS strain 5448 with ska(type-2b) caused a reduction in virulence while exchanging ska(type-2a) with ska(type-1) into 5448 produced an increase in virulence when using a mouse model of invasive disease. These findings suggest that streptokinase variants produced by GAS isolates utilize distinct plasminogen activation pathways, which directly affects the pathogenesis of this organism.

  8. Molecular and genomic characterization of pathogenic traits of group A Streptococcus pyogenes

    PubMed Central

    HAMADA, Shigeyuki; KAWABATA, Shigetada; NAKAGAWA, Ichiro

    2015-01-01

    Group A streptococcus (GAS) or Streptococcus pyogenes causes various diseases ranging from self-limiting sore throat to deadly invasive diseases. The genome size of GAS is 1.85–1.9 Mb, and genomic rearrangement has been demonstrated. GAS possesses various surface-associated substances such as hyaluronic capsule, M proteins, and fibronectin/laminin/immunoglobulin-binding proteins. These are related to the virulence and play multifaceted and mutually reflected roles in the pathogenesis of GAS infections. Invasion of GAS into epithelial cells and deeper tissues provokes immune and non-immune defense or inflammatory responses including the recruitment of neutrophils, macrophages, and dendritic cells in hosts. GAS frequently evades host defense mechanisms by using its virulence factors. Extracellular products of GAS may perturb cellular and subcellular functions and degrade tissues enzymatically, which leads to the aggravation of local and/or systemic disorders in the host. In this review, we summarize some important cellular and extracellular substances that may affect pathogenic processes during GAS infections, and the host responses to these. PMID:26666305

  9. Molecular characterization of Streptococcus pyogenes group A isolates from a tertiary hospital in Lebanon.

    PubMed

    Karaky, Nathalie M; Araj, George F; Tokajian, Sima T

    2014-09-01

    Streptococcus pyogenes [Group A Streptococcus (GAS)] is one of the most important human pathogens, responsible for numerous diseases with diverse clinical manifestations. As the epidemiology of GAS infections evolves, a rapid and reliable characterization of the isolates remains essential for epidemiological analysis and infection control. This study investigated the epidemiological patterns and genetic characteristics of 150 GAS isolates from a tertiary hospital in Lebanon by emm typing, superantigens (SAgs) detection, PFGE and antibiotic profiling. The results revealed 41 distinct emm types, the most prevalent of which were emm89 (16 %), emm12 (10 %), emm2 (9 %) and emm1 (8 %). Testing for the presence of superantigens showed that speB (87 %), ssa (36 %) and speG (30 %) were predominant. PFGE detected 39 pulsotypes when a similarity cut-off value of 80 % was implemented. Antibiotic-susceptibility testing against seven different classes of antibiotics showed that 9 % of the isolates were resistant to clindamycin, 23 % were resistant to erythromycin and 4 % showed the macrolide-lincosamide-streptogramin B (MLSB) phenotype. The emergence of tetracycline-resistant strains (37 %) was high when compared with previous reports from Lebanon. This study provided comprehensive evidence of the epidemiology of GAS in Lebanon, highlighting the association between emm types and toxin genes, and providing valuable information about the origin and dissemination of this pathogen. © 2014 The Authors.

  10. Essential Genes in the Core Genome of the Human Pathogen Streptococcus pyogenes

    PubMed Central

    Le Breton, Yoann; Belew, Ashton T.; Valdes, Kayla M.; Islam, Emrul; Curry, Patrick; Tettelin, Hervé; Shirtliff, Mark E.; El-Sayed, Najib M.; McIver, Kevin S.

    2015-01-01

    Streptococcus pyogenes (Group A Streptococcus, GAS) remains a major public health burden worldwide, infecting over 750 million people leading to over 500,000 deaths annually. GAS pathogenesis is complex, involving genetically distinct GAS strains and multiple infection sites. To overcome fastidious genetic manipulations and accelerate pathogenesis investigations in GAS, we developed a mariner-based system (Krmit) for en masse monitoring of complex mutant pools by transposon sequencing (Tn-seq). Highly saturated transposant libraries (Krmit insertions in ca. every 25 nucleotides) were generated in two distinct GAS clinical isolates, a serotype M1T1 invasive strain 5448 and a nephritogenic serotype M49 strain NZ131, and analyzed using a Bayesian statistical model to predict GAS essential genes, identifying sets of 227 and 241 of those genes in 5448 and NZ131, respectively. A large proportion of GAS essential genes corresponded to key cellular processes and metabolic pathways, and 177 were found conserved within the GAS core genome established from 20 available GAS genomes. Selected essential genes were validated using conditional-expression mutants. Finally, comparison to previous essentiality analyses in S. sanguinis and S. pneumoniae revealed significant overlaps, providing valuable insights for the development of new antimicrobials to treat infections by GAS and other pathogenic streptococci. PMID:25996237

  11. The Regulatory Small RNA MarS Supports Virulence of Streptococcus pyogenes.

    PubMed

    Pappesch, Roberto; Warnke, Philipp; Mikkat, Stefan; Normann, Jana; Wisniewska-Kucper, Aleksandra; Huschka, Franziska; Wittmann, Maja; Khani, Afsaneh; Schwengers, Oliver; Oehmcke-Hecht, Sonja; Hain, Torsten; Kreikemeyer, Bernd; Patenge, Nadja

    2017-09-25

    Small regulatory RNAs (sRNAs) play a role in the control of bacterial virulence gene expression. In this study, we investigated an sRNA that was identified in Streptococcus pyogenes (group A Streptococcus, GAS) but is conserved throughout various streptococci. In a deletion strain, expression of mga, the gene encoding the multiple virulence gene regulator, was reduced. Accordingly, transcript and proteome analyses revealed decreased expression of several Mga-activated genes. Therefore, and because the sRNA was shown to interact with the 5' UTR of the mga transcript in a gel-shift assay, we designated it MarS for m ga-activating regulatory sRNA. Down-regulation of important virulence factors, including the antiphagocytic M-protein, led to increased susceptibility of the deletion strain to phagocytosis and reduced adherence to human keratinocytes. In a mouse infection model, the marS deletion mutant showed reduced dissemination to the liver, kidney, and spleen. Additionally, deletion of marS led to increased tolerance towards oxidative stress. Our in vitro and in vivo results indicate a modulating effect of MarS on virulence gene expression and on the pathogenic potential of GAS.

  12. Spontaneous mutations in Streptococcus pyogenes isolates from streptococcal toxic shock syndrome patients play roles in virulence

    PubMed Central

    Ikebe, Tadayoshi; Matsumura, Takayuki; Nihonmatsu, Hisako; Ohya, Hitomi; Okuno, Rumi; Mitsui, Chieko; Kawahara, Ryuji; Kameyama, Mitsuhiro; Sasaki, Mari; Shimada, Naomi; Ato, Manabu; Ohnishi, Makoto

    2016-01-01

    Streptococcus pyogenes (group A Streptococcus; GAS) is a widespread human pathogen and causes streptococcal toxic shock syndrome (STSS). STSS isolates have been previously shown to have high frequency mutations in the csrS/csrR (covS/covR) and/or rgg (ropB) genes, which are negative regulators of virulence. However, these mutations were found at somewhat low frequencies in emm1-genotyped isolates, the most prevalent STSS genotype. In this study, we sought to detect causal mutations of enhanced virulence in emm1 isolates lacking mutation(s) in the csrS/csrR and rgg genes. Three mutations associated with elevated virulence were found in the sic (a virulence gene) promoter, the csrR promoter, and the rocA gene (a csrR positive regulator). In vivo contribution of the sic promoter and rocA mutations to pathogenicity and lethality was confirmed in a GAS mouse model. Frequency of the sic promoter mutation was significantly higher in STSS emm1 isolates than in non-invasive STSS isolates; the rocA gene mutation frequency was not significantly different among STSS and non-STSS isolates. STSS emm1 isolates possessed a high frequency mutation in the sic promoter. Thus, this mutation may play a role in the dynamics of virulence and STSS pathogenesis. PMID:27349341

  13. PepO, a CovRS-controlled endopeptidase, disrupts Streptococcus pyogenes quorum sensing.

    PubMed

    Wilkening, Reid V; Chang, Jennifer C; Federle, Michael J

    2016-01-01

    Group A Streptococcus (GAS, Streptococcus pyogenes) is a human-restricted pathogen with a capacity to both colonize asymptomatically and cause illnesses ranging from pharyngitis to necrotizing fasciitis. An understanding of how and when GAS switches between genetic programs governing these different lifestyles has remained an enduring mystery and likely requires carefully tuned environmental sensors to activate and silence genetic schemes when appropriate. Herein, we describe the relationship between the Control of Virulence (CovRS, CsrRS) two-component system and the Rgg2/3 quorum-sensing pathway. We demonstrate that responses of CovRS to the stress signals Mg(2+) and a fragment of the antimicrobial peptide LL-37 result in modulated activity of pheromone signaling of the Rgg2/3 pathway through a means of proteolysis of SHP peptide pheromones. This degradation is mediated by the cytoplasmic endopeptidase PepO, which is the first identified enzymatic silencer of an RRNPP-type quorum-sensing pathway. These results suggest that under conditions in which the virulence potential of GAS is elevated (i.e. enhanced virulence gene expression), cellular responses mediated by the Rgg2/3 pathway are abrogated and allow individuals to escape from group behavior. These results also indicate that Rgg2/3 signaling is instead functional during non-virulent GAS lifestyles. © 2015 John Wiley & Sons Ltd.

  14. PepO, a CovRS-controlled endopeptidase, disrupts Streptococcus pyogenes quorum sensing

    PubMed Central

    Wilkening, Reid V.; Chang, Jennifer C.; Federle, Michael J.

    2016-01-01

    Summary Group A Streptococcus (GAS, Streptococcus pyogenes) is a human-restricted pathogen with a capacity to both colonize asymptomatically and cause illnesses ranging from pharyngitis to necrotizing fasciitis. An understanding of how and when GAS switches between genetic programs governing these different lifestyles has remained an enduring mystery and likely requires carefully tuned environmental sensors to activate and silence genetic schemes when appropriate. Herein, we describe the relationship between the Control of Virulence (CovRS, CsrRS) two-component system and the Rgg2/3 quorum-sensing pathway. We demonstrate that responses of CovRS to the stress signals Mg2+ and a fragment of the antimicrobial peptide LL-37 result in modulated activity of pheromone signaling of the Rgg2/3 pathway through a means of proteolysis of SHP peptide pheromones. This degradation is mediated by the cytoplasmic endopeptidase PepO, which is the first identified enzymatic silencer of an RRNPP-type quorum-sensing pathway. These results suggest that under conditions in which the virulence potential of GAS is elevated (i.e. enhanced virulence gene expression), cellular responses mediated by the Rgg2/3 pathway are abrogated and allow individuals to escape from group behavior. These results also indicate that Rgg2/3 signaling is instead functional during non-virulent GAS lifestyles. PMID:26418177

  15. Bactericidal activity of moxifloxacin compared to grepafloxacin and clarithromycin against Streptococcus pneumoniae and Streptococcus pyogenes investigated using an in vitro pharmacodynamic model.

    PubMed

    Esposito, S; Noviello, S; Ianniello, F

    2000-12-01

    The aim of the present investigation was to study and compare the killing activity of two new fluoroquinolone compounds, moxifloxacin and grepafloxacin, and a new generation macrolide, clarithromycin, against three clinical isolates of Streptococcus pneumoniae (penicillin-susceptible, -intermediate and -resistant) and two Streptococcus pyogenes (erythromycin-susceptible and -resistant) strains by simulating their human pharmacokinetics in a pharmacodynamic model. Results were achieved by measuring the reduction in viable bacterial count during the 24-h experimental period. All three antimicrobials led to a continuous reduction in the bacterial counts of penicillin-susceptible S. pneumoniae and erythromycin-susceptible S. pyogenes strains, the maximal reduction observed after 8-10 hours being 5-6 logs for moxifloxacin and 3 logs for grepafloxacin; clarithromycin exhibited a similar reduction of 5 logs only after 24 h. No regrowth was observed for any strain after 24 h with any of the antibiotics. The bactericidal activity of both the fluoroquinolones was not affected by penicillin resistance of S. pneumoniae and erythromycin resistance of S. pyogenes. In contrast, clarithromycin was not able to reduce the bacterial count of penicillin-resistant S. pneumoniae and erythromycin-resistant S. pyogenes strains. Moxifloxacin exhibited, within 24 h, higher and faster bactericidal activity than grepafloxacin and clarithromycin against S. pneumoniae, and was not affected by penicillin resistance. These results suggest that moxifloxacin is a promising new agent for treatment of streptococcal infections.

  16. Streptococcus pyogenes Pharyngeal Isolates with Reduced Susceptibility to Ciprofloxacin in Spain: Mechanisms of Resistance and Clonal Diversity

    PubMed Central

    Albertí, Sebastián; Cortés, Guadalupe; García-Rey, Cesar; Rubio, Carmen; Baquero, Fernando; García-Rodríguez, José Ángel; Bouza, Emilio; Aguilar, Lorenzo

    2005-01-01

    A survey of emm gene sequences and an analysis of the pulsed-field electrophoretic profiles of 30 Streptococcus pyogenes isolates with reduced susceptibilities to ciprofloxacin detected the prevalence of isolates with emm type 6 and considerable genetic diversity among isolates. The mechanism of ciprofloxacin resistance in these isolates was based on point mutations in topoisomerase IV subunit C encoded by parC, mainly replacement of serine-79 by alanine. PMID:15616324

  17. Rapid Development of Brain Abscess Caused by Streptococcus Pyogenes Following Penetrating Skull Injury via the Ethmoidal Sinus and Lamina Cribrosa

    PubMed Central

    Aydin, Gerilmez; Cömert, Serhat; Altinors, Nur

    2010-01-01

    Objective Streptococcus pyogenes is a beta-hemolytic bacterium that belongs to Lancefield serogroup A, also known as group A streptococci (GAS). There have been five reported case in terms of PubMed-based search but no reported case of brain abscess caused by Streptococcus pyogenes as a result of penetrating skull injury. We present a patient who suffered from penetrating skull injury that resulted in a brain abscess caused by Streptococcus pyogenes. Methods The patient was a 12-year-old boy who fell down from his bicycle while cycling and ran into a tree. A wooden stick penetrated his skin below the right lower eyelid and advanced to the cranium. He lost consciousness on the fifth day of the incident and his body temperature was measured as 40℃. While being admitted to our hospital, a cranial computed tomography revealed a frontal cystic mass with a perilesional hypodense zone of edema. There was no capsule formation around the lesion after intravenous contrast injection. Paranasal CT showed a bone defect located between the ethmoidal sinus and lamina cribrosa. Results Bifrontal craniotomy was performed. The abscess located at the left frontal lobe was drained and the bone defect was repaired. Conclusion Any penetrating lesion showing a connection between the lamina cribrosa and ethmoidal sinus may result in brain abscess caused by Streptococcus pyogenes. These patients should be treated urgently to repair the defect and drain the abscess with appropriate antibiotic therapy started due to the fulminant course of the brain abscess caused by this microorganism. PMID:20717517

  18. Salivaricin G32, a Homolog of the Prototype Streptococcus pyogenes Nisin-Like Lantibiotic SA-FF22, Produced by the Commensal Species Streptococcus salivarius

    PubMed Central

    Wescombe, Philip A.; Dyet, Kristin H.; Dierksen, Karen P.; Power, Daniel A.; Jack, Ralph W.; Burton, Jeremy P.; Inglis, Megan A.; Wescombe, Anna L.; Tagg, John R.

    2012-01-01

    Salivaricin G32, a 2667 Da novel member of the SA-FF22 cluster of lantibiotics, has been purified and characterized from Streptococcus salivarius strain G32. The inhibitory peptide differs from the Streptococcus pyogenes—produced SA-FF22 in the absence of lysine in position 2. The salivaricin G32 locus was widely distributed in BLIS-producing S. salivarius, with 6 (23%) of 26 strains PCR-positive for the structural gene, slnA. As for most other lantibiotics produced by S. salivarius, the salivaricin G32 locus can be megaplasmid encoded. Another member of the SA-FF22 family was detected in two Streptococcus dysgalactiae of bovine origin, an observation supportive of widespread distribution of this lantibiotic within the genus Streptococcus. Since the inhibitory spectrum of salivaricin G32 includes Streptococcus pyogenes, its production by S. salivarius, either as a member of the normal oral microflora or as a commercial probiotic, could serve to enhance protection of the human host against S. pyogenes infection. PMID:22567013

  19. Identification of a two-component Class IIb bacteriocin in Streptococcus pyogenes by recombinase-based in vivo expression technology.

    PubMed

    Armstrong, Brent D; Herfst, Christine A; Tonial, Nicholas C; Wakabayashi, Adrienne T; Zeppa, Joseph J; McCormick, John K

    2016-11-03

    Streptococcus pyogenes is a globally prominent bacterial pathogen that exhibits strict tropism for the human host, yet bacterial factors responsible for the ability of S. pyogenes to compete within this limited biological niche are not well understood. Using an engineered recombinase-based in vivo expression technology (RIVET) system, we identified an in vivo-induced promoter region upstream of a predicted Class IIb bacteriocin system in the M18 serotype S. pyogenes strain MGAS8232. This promoter element was not active under in vitro laboratory conditions, but was highly induced within the mouse nasopharynx. Recombinant expression of the predicted mature S. pyogenes bacteriocin peptides (designated SpbM and SpbN) revealed that both peptides were required for antimicrobial activity. Using a gain of function experiment in Lactococcus lactis, we further demonstrated S. pyogenes immunity function is encoded downstream of spbN. These data highlight the importance of bacterial gene regulation within appropriate environments to help understand mechanisms of niche adaptation by bacterial pathogens.

  20. Identification of a two-component Class IIb bacteriocin in Streptococcus pyogenes by recombinase-based in vivo expression technology

    PubMed Central

    Armstrong, Brent D.; Herfst, Christine A.; Tonial, Nicholas C.; Wakabayashi, Adrienne T.; Zeppa, Joseph J.; McCormick, John K.

    2016-01-01

    Streptococcus pyogenes is a globally prominent bacterial pathogen that exhibits strict tropism for the human host, yet bacterial factors responsible for the ability of S. pyogenes to compete within this limited biological niche are not well understood. Using an engineered recombinase-based in vivo expression technology (RIVET) system, we identified an in vivo-induced promoter region upstream of a predicted Class IIb bacteriocin system in the M18 serotype S. pyogenes strain MGAS8232. This promoter element was not active under in vitro laboratory conditions, but was highly induced within the mouse nasopharynx. Recombinant expression of the predicted mature S. pyogenes bacteriocin peptides (designated SpbM and SpbN) revealed that both peptides were required for antimicrobial activity. Using a gain of function experiment in Lactococcus lactis, we further demonstrated S. pyogenes immunity function is encoded downstream of spbN. These data highlight the importance of bacterial gene regulation within appropriate environments to help understand mechanisms of niche adaptation by bacterial pathogens. PMID:27808235

  1. IL-2 Expression and T lymphocyte Phenotyping in Young Children Suffering from Upper Respiratory Tract Infection with Streptococcus Pyogenes

    PubMed Central

    Guadalupe Ramirez-Valles, Eda; Dayali Gutierrez-Martinez, Verónica; Cervantes-Flores, Maribel; Ruiz-Baca, Estela; Alvarado-Esquivel, Cosme

    2016-01-01

    T cells are components of adaptive immunity and are involved in the resolution of respiratory infections, which are a major cause of morbidity and mortality in young children worldwide. Activation and differentiation of T cells is given mostly by the cytokine IL-2. This study aimed to determine the phenotype of T cells and IL-2 expression in children suffering from upper respiratory tract infection with Streptococcus pyogenes (S. pyogenes). For this purpose, IL-2 expression at its gene and protein levels and quantitation of CD4+ and CD8+ T lymphocytes were assessed in children aged 0-5 years old suffering from upper respiratory tract infection with S. pyogenes and healthy children of the same age. Children with S. pyogenes infection had a higher expression of IL-2 gene and a lower level of this cytokine expression at protein level than healthy children. The numbers of CD4+ T lymphocytes were similar among the groups. In contrast, difference in the numbers of CD8+ T lymphocytes among the groups was found. We conclude that infections by S. pyogenes in young children lead to an increased expression of IL-2 mRNA. PMID:27493590

  2. First Streptococcus pyogenes signature-tagged mutagenesis screen identifies novel virulence determinants.

    PubMed

    Kizy, Anne E; Neely, Melody N

    2009-05-01

    The virulence of bacterial pathogens is a complex process that requires the dynamic expression of many genes for the pathogens to invade and circumvent host defenses, as well as to proliferate in vivo. In this study, we employed a large-scale screen, signature-tagged mutagenesis (STM), to identify Streptococcus pyogenes virulence genes important for pathogenesis within the host. Approximately 1,200 STM mutants were created and screened using the zebrafish infectious disease model. The transposon insertion site was identified for 29 of the 150 mutants that were considered attenuated for virulence. Previously reported streptococcal virulence genes, such as mga, hasA, amrA, smeZ, and two genes in the sil locus, were identified, confirming the utility of the model for revealing genes important for virulence. Multiple genes not previously implicated in virulence were also identified, including genes encoding putative transporters, hypothetical cytosolic proteins, and macrolide efflux pumps. The STM mutant strains display various levels of attenuation, and multiple separate insertions were identified in either the same gene or the same locus, suggesting that these factors are important for this type of acute, invasive infection. We further examined two such genes, silB and silC of a putative quorum-sensing regulon, and determined that they are significant virulence factors in our model of necrotizing fasciitis. sil locus promoter expression was examined under various in vitro conditions, as well as in zebrafish tissues, and was found to be differentially induced. This study was a unique investigation of S. pyogenes factors required for successful invasive infection.

  3. A Novel Role for Pro-Coagulant Microvesicles in the Early Host Defense against Streptococcus pyogenes

    PubMed Central

    Oehmcke, Sonja; Westman, Johannes; Malmström, Johan; Mörgelin, Matthias; Olin, Anders I.; Kreikemeyer, Bernd; Herwald, Heiko

    2013-01-01

    Previous studies have shown that stimulation of whole blood or peripheral blood mononuclear cells with bacterial virulence factors results in the sequestration of pro-coagulant microvesicles (MVs). These particles explore their clotting activity via the extrinsic and intrinsic pathway of coagulation; however, their pathophysiological role in infectious diseases remains enigmatic. Here we describe that the interaction of pro-coagulant MVs with bacteria of the species Streptococcus pyogenes is part of the early immune response to the invading pathogen. As shown by negative staining electron microscopy and clotting assays, pro-coagulant MVs bind in the presence of plasma to the bacterial surface. Fibrinogen was identified as a linker that, through binding to the M1 protein of S. pyogenes, allows the opsonization of the bacteria by MVs. Surface plasmon resonance analysis revealed a strong interaction between pro-coagulant MVs and fibrinogen with a KD value in the nanomolar range. When performing a mass-spectrometry-based strategy to determine the protein quantity, a significant up-regulation of the fibrinogen-binding integrins CD18 and CD11b on pro-coagulant MVs was recorded. Finally we show that plasma clots induced by pro-coagulant MVs are able to prevent bacterial dissemination and possess antimicrobial activity. These findings were confirmed by in vivo experiments, as local treatment with pro-coagulant MVs dampens bacterial spreading to other organs and improved survival in an invasive streptococcal mouse model of infection. Taken together, our data implicate that pro-coagulant MVs play an important role in the early response of the innate immune system in infectious diseases. PMID:23935504

  4. The pyogenic potential of the different Streptococcus anginosus group bacterial species: retrospective cohort study.

    PubMed

    Kobo, O; Nikola, S; Geffen, Y; Paul, M

    2017-08-14

    Streptococcus anginosus Group (SAG) bacteria are common causes of pyogenic infections (PIs). We examined the association between SAG species and the presence of a PI through a retrospective, observational, cohort study, between the years 2009 and 2015. All adults with clinically significant SAG infections in one hospital in Israel were assessed for association between SAG species and the presence of a PI defined as an abscess, empyema, or deep/organ space surgical site infection. Risk factors for PI were assessed using multivariate backward stepwise logistic regression analysis. We identified 263 patients with significant SAG infections, 182 (69%) of which were caused by S. anginosus, 45 (17·1%) by S treptococcus constellatus and 36 (13·7%) by S treptococcus intermedius. The mean age of the patients was 56·8 ± 19·1 years. PIs were identified among 160 (60%) of the patients and were mostly non-bacteraemic (147/160, 91·8%), while most non-PI patients had bacteraemia (70/103, 68%). S. anginosus and S. constellatus were associated with a significantly lower incidence of PI than S. intermedius, OR 0·18 (95% CI 0·06-0·53) and 0·14 (0·04-0·48), respectively. Patients with PI were younger and, in general, had less co-morbidities. S. intermedius was associated with pyogenic non-bacteraemic infections, while S. anginosus and S. constellatus were associated with bacteraemia with no abscess or empyema formation. These data may indicate differences in virulence mechanisms of these SAG bacteria.

  5. Isopeptide bonds block the mechanical extension of pili in pathogenic Streptococcus pyogenes.

    PubMed

    Alegre-Cebollada, Jorge; Badilla, Carmen L; Fernández, Julio M

    2010-04-09

    In the early stages of an infection, pathogenic bacteria use long fibrous structures known as pili as adhesive anchors for attachment to the host cells. These structures also play key roles in colony and biofilm formation. In all those processes, pili must withstand large mechanical forces. The pili of the nasty gram-positive human pathogen Streptococcus pyogenes are assembled as single, micrometer long tandem modular proteins of covalently linked repeats of pilin proteins. Here we use single molecule force spectroscopy techniques to study the mechanical properties of the major pilin Spy0128. In our studies, we engineer polyproteins containing repeats of Spy0128 flanked by the well characterized I27 protein which provides an unambiguous mechanical fingerprint. We find that Spy0128 is an inextensible protein, even when pulled at forces of up to 800 pN. We also found that this remarkable mechanical resilience, unique among the modular proteins studied to date, results from the strategically located intramolecular isopeptide bonds recently identified in the x-ray structure of Spy0128. Removal of the isopeptide bonds by mutagenesis readily allowed Spy0128 domains to unfold and extend, albeit at relatively high forces of 172 pN (N-terminal domain) or 250 pN (C-terminal domain). Our results show that in contrast to the elastic roles played by large tandem modular proteins such as titin and fibronectin, the giant pili of S. pyogenes evolved to abrogate mechanical extensibility, a property that may be crucial in the pathogenesis of this most virulent bacterium and, therefore, become the target of new therapeutic approaches against its infections.

  6. Antagonistic Rgg regulators mediate quorum sensing via competitive DNA binding in Streptococcus pyogenes.

    PubMed

    Lasarre, Breah; Aggarwal, Chaitanya; Federle, Michael J

    2013-01-02

    Recent studies have established the fact that multiple members of the Rgg family of transcriptional regulators serve as key components of quorum sensing (QS) pathways that utilize peptides as intercellular signaling molecules. We previously described a novel QS system in Streptococcus pyogenes which utilizes two Rgg-family regulators (Rgg2 and Rgg3) that respond to neighboring signaling peptides (SHP2 and SHP3) to control gene expression and biofilm formation. We have shown that Rgg2 is a transcriptional activator of target genes, whereas Rgg3 represses expression of these genes, and that SHPs function to activate the QS system. The mechanisms by which Rgg proteins regulate both QS-dependent and QS-independent processes remain poorly defined; thus, we sought to further elucidate how Rgg2 and Rgg3 mediate gene regulation. Here we provide evidence that S. pyogenes employs a unique mechanism of direct competition between the antagonistic, peptide-responsive proteins Rgg2 and Rgg3 for binding at target promoters. The highly conserved, shared binding sites for Rgg2 and Rgg3 are located proximal to the -35 nucleotide in the target promoters, and the direct competition between the two regulators results in concentration-dependent, exclusive occupation of the target promoters that can be skewed in favor of Rgg2 in vitro by the presence of SHP. These results suggest that exclusionary binding of target promoters by Rgg3 may prevent Rgg2 binding under SHP-limiting conditions, thereby preventing premature induction of the quorum sensing circuit. Rgg-family transcriptional regulators are widespread among low-G+C Gram-positive bacteria and in many cases contribute to bacterial physiology and virulence. Only recently was it discovered that several Rgg proteins function in cell-to-cell communication (quorum sensing [QS]) via direct interaction with signaling peptides. The mechanism(s) by which Rgg proteins mediate regulation is poorly understood, and further insight into Rgg

  7. An association between peptidoglycan synthesis and organization of the Streptococcus pyogenes ExPortal.

    PubMed

    Vega, Luis Alberto; Port, Gary C; Caparon, Michael G

    2013-09-24

    The ExPortal of Streptococcus pyogenes is a focal microdomain of the cytoplasmic membrane that clusters the translocons of the general secretory pathway with accessory factors to facilitate the maturation of secreted polypeptides. While it is known that the ExPortal is enriched in anionic lipids, the mechanisms that organize the ExPortal are poorly understood. In the present study, we examined the role of the cell wall in organizing and maintaining the ExPortal. Removal of the cell wall resulted in a loss of ExPortal focal integrity accompanied by the circumferential redistribution of ExPortal lipid and protein components. A similar loss occurred upon treatment with gallidermin, a nonpermeabilizing lantibiotic that targets the lipid II precursor of peptidoglycan synthesis, and this treatment disrupted the secretion of several ExPortal substrates. Furthermore, several enzymes involved in the membrane-associated steps of lipid II synthesis, including MraY and MurN, were found to localize to a single discrete focus in the membrane that was coincident with the focal location of the secretory translocons and the anionic lipid microdomain. These data suggest that the ExPortal is associated with the site of peptidoglycan precursor synthesis and that peptidoglycan biogenesis influences ExPortal organization. These data add to an emerging literature indicating that cell wall biogenesis, cell division, and protein secretion are spatially coorganized processes. Since Gram-positive bacteria lack a periplasmic space, they lack a protected compartment to spatially coordinate interaction between newly secreted proteins and the factors required to process them. This represents a significant problem for pathogens that depend on the secretion of toxins and cell wall-associated adhesins to cause disease. Streptococci solve this dilemma by restricting secretion and processing factors to a defined region of the membrane. However, the mechanisms that promote restriction are not

  8. Identification and characterization of two temperature-induced surface-associated proteins of Streptococcus suis with high homologies to members of the Arginine Deiminase system of Streptococcus pyogenes.

    PubMed

    Winterhoff, Nora; Goethe, Ralph; Gruening, Petra; Rohde, Manfred; Kalisz, Henryk; Smith, Hilde E; Valentin-Weigand, Peter

    2002-12-01

    The present study was performed to identify stress-induced putative virulence proteins of Streptococcus suis. For this, protein expression patterns of streptococci grown at 32, 37, and 42 degrees C were compared by one- and two-dimensional gel electrophoresis. Temperature shifts from 32 and 37 to 42 degrees C induced expression of two cell wall-associated proteins with apparent molecular masses of approximately 47 and 53 kDa. Amino-terminal sequence analysis of the two proteins indicated homologies of the 47-kDa protein with an ornithine carbamoyltransferase (OCT) from Streptococcus pyogenes and of the 53-kDa protein with the streptococcal acid glycoprotein (SAGP) from S. pyogenes, an arginine deiminase (AD) recently proposed as a putative virulence factor. Cloning and sequencing the genes encoding the putative OCT and AD of S. suis, octS and adiS, respectively, revealed that they had 81.2 (octS) and 80.2% (adiS) identity with the respective genes of S. pyogenes. Both genes belong to the AD system, also found in other bacteria. Southern hybridization analysis demonstrated the presence of the adiS gene in all 42 serotype 2 and 9 S. suis strains tested. In 9 of these 42 strains, selected randomly, we confirmed expression of the AdiS protein, homologous to SAGP, by immunoblot analysis using a specific antiserum against the SAGP of S. pyogenes. In all strains AD activity was detected. Furthermore, by immunoelectron microscopy using the anti-S. pyogenes SAGP antiserum we were able to demonstrate that the AdiS protein is expressed on the streptococcal surface in association with the capsular polysaccharides but is not coexpressed with them.

  9. Identification and Characterization of Two Temperature-Induced Surface-Associated Proteins of Streptococcus suis with High Homologies to Members of the Arginine Deiminase System of Streptococcus pyogenes

    PubMed Central

    Winterhoff, Nora; Goethe, Ralph; Gruening, Petra; Rohde, Manfred; Kalisz, Henryk; Smith, Hilde E.; Valentin-Weigand, Peter

    2002-01-01

    The present study was performed to identify stress-induced putative virulence proteins of Streptococcus suis. For this, protein expression patterns of streptococci grown at 32, 37, and 42°C were compared by one- and two-dimensional gel electrophoresis. Temperature shifts from 32 and 37 to 42°C induced expression of two cell wall-associated proteins with apparent molecular masses of approximately 47 and 53 kDa. Amino-terminal sequence analysis of the two proteins indicated homologies of the 47-kDa protein with an ornithine carbamoyltransferase (OCT) from Streptococcus pyogenes and of the 53-kDa protein with the streptococcal acid glycoprotein (SAGP) from S. pyogenes, an arginine deiminase (AD) recently proposed as a putative virulence factor. Cloning and sequencing the genes encoding the putative OCT and AD of S. suis, octS and adiS, respectively, revealed that they had 81.2 (octS) and 80.2% (adiS) identity with the respective genes of S. pyogenes. Both genes belong to the AD system, also found in other bacteria. Southern hybridization analysis demonstrated the presence of the adiS gene in all 42 serotype 2 and 9 S. suis strains tested. In 9 of these 42 strains, selected randomly, we confirmed expression of the AdiS protein, homologous to SAGP, by immunoblot analysis using a specific antiserum against the SAGP of S. pyogenes. In all strains AD activity was detected. Furthermore, by immunoelectron microscopy using the anti-S. pyogenes SAGP antiserum we were able to demonstrate that the AdiS protein is expressed on the streptococcal surface in association with the capsular polysaccharides but is not coexpressed with them. PMID:12446626

  10. The microbiology of impetigo in indigenous children: associations between Streptococcus pyogenes, Staphylococcus aureus, scabies, and nasal carriage.

    PubMed

    Bowen, Asha C; Tong, Steven Y C; Chatfield, Mark D; Carapetis, Jonathan R

    2014-12-31

    Impetigo is caused by both Streptococcus pyogenes and Staphylococcus aureus; the relative contributions of each have been reported to fluctuate with time and region. While S. aureus is reportedly on the increase in most industrialised settings, S. pyogenes is still thought to drive impetigo in endemic, tropical regions. However, few studies have utilised high quality microbiological culture methods to confirm this assumption. We report the prevalence and antimicrobial resistance of impetigo pathogens recovered in a randomised, controlled trial of impetigo treatment conducted in remote Indigenous communities of northern Australia. Each child had one or two sores, and the anterior nares, swabbed. All swabs were transported in skim milk tryptone glucose glycogen broth and frozen at -70°C, until plated on horse blood agar. S. aureus and S. pyogenes were confirmed with latex agglutination. From 508 children, we collected 872 swabs of sores and 504 swabs from the anterior nares prior to commencement of antibiotic therapy. S. pyogenes and S. aureus were identified together in 503/872 (58%) of sores; with an additional 207/872 (24%) sores having S. pyogenes and 81/872 (9%) S. aureus, in isolation. Skin sore swabs taken during episodes with a concurrent diagnosis of scabies were more likely to culture S. pyogenes (OR 2.2, 95% CI 1.1 - 4.4, p = 0.03). Eighteen percent of children had nasal carriage of skin pathogens. There was no association between the presence of S. aureus in the nose and skin. Methicillin-resistance was detected in 15% of children who cultured S. aureus from either a sore or their nose. There was no association found between the severity of impetigo and the detection of a skin pathogen. S. pyogenes remains the principal pathogen in tropical impetigo; the relatively high contribution of S. aureus as a co-pathogen has also been confirmed. Children with scabies were more likely to have S. pyogenes detected. While clearance of S. pyogenes is the key

  11. Biochemical and biological activity of arginine deiminase from Streptococcus pyogenes M22.

    PubMed

    Starikova, Eleonora A; Sokolov, Alexey V; Vlasenko, Anna Yu; Burova, Larisa A; Freidlin, Irina S; Vasilyev, Vadim B

    2016-04-01

    Streptococcus pyogenes (group A Streptococcus; GAS) is an important gram-positive extracellular bacterial pathogen responsible for a number of suppurative infections. This micro-organism has developed complex virulence mechanisms to avoid the host's defenses. We have previously reported that SDSC from GAS type M22 causes endothelial-cell dysfunction, and inhibits cell adhesion, migration, metabolism, and proliferation in a dose-dependent manner, without affecting cell viability. This work aimed to isolate and characterize a component from GAS type M22 supernatant that suppresses the proliferation of endothelial cells (EA.hy926). In the process of isolating a protein possessing antiproliferative activity we identified arginine deiminase (AD). Further study showed that this enzyme is most active at pH 6.8. Calculating Km and Vmax gave the values of 0.67 mmol·L(-1) and 42 s(-1), respectively. A distinctive feature of AD purified from GAS type M22 is that its optimum activity and the maximal rate of the catalytic process is close to neutral pH by comparison with enzymes from other micro-organisms. AD from GAS type M22 suppressed the proliferative activity of endothelial cells in a dose-dependent mode. At the same time, in the presence of AD, the proportion of cells in G0/G1 phase increased. When l-Arg was added at increasing concentrations to the culture medium containing AD (3 μg·mL(-1)), the enzyme's capacity to inhibit cell proliferation became partially depressed. The proportion of cells in phases S/G2 increased concomitantly, although the cells did not fully recover their proliferation activity. This suggests that AD from GAS type M22 has potential for the suppression of excessive cell proliferation.

  12. Streptococcal 5'-Nucleotidase A (S5nA), a Novel Streptococcus pyogenes Virulence Factor That Facilitates Immune Evasion.

    PubMed

    Zheng, Lisa; Khemlani, Adrina; Lorenz, Natalie; Loh, Jacelyn M S; Langley, Ries J; Proft, Thomas

    2015-12-25

    Streptococcus pyogenes is an important human pathogen that causes a wide range of diseases. Using bioinformatics analysis of the complete S. pyogenes strain SF370 genome, we have identified a novel S. pyogenes virulence factor, which we termed streptococcal 5'-nucleotidase A (S5nA). A recombinant form of S5nA hydrolyzed AMP and ADP, but not ATP, to generate the immunomodulatory molecule adenosine. Michaelis-Menten kinetics revealed a Km of 169 μm and a Vmax of 7550 nmol/mg/min for the substrate AMP. Furthermore, recombinant S5nA acted synergistically with S. pyogenes nuclease A to generate macrophage-toxic deoxyadenosine from DNA. The enzyme showed optimal activity between pH 5 and pH 6.5 and between 37 and 47 °C. Like other 5'-nucleotidases, S5nA requires divalent cations and was active in the presence of Mg(2+), Ca(2+), or Mn(2+). However, Zn(2+) inhibited the enzymatic activity. Structural modeling combined with mutational analysis revealed a highly conserved catalytic dyad as well as conserved substrate and cation-binding sites. Recombinant S5nA significantly increased the survival of the non-pathogenic bacterium Lactococcus lactis during a human whole blood killing assay in a dose-dependent manner, suggesting a role as an S. pyogenes virulence factor. In conclusion, we have identified a novel S. pyogenes enzyme with 5'-nucleotidase activity and immune evasion properties. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Presence of a Prophage Determines Temperature-Dependent Capsule Production in Streptococcus pyogenes

    PubMed Central

    Brown, Leslie; Kim, Jeong-Ho; Cho, Kyu Hong

    2016-01-01

    A hyaluronic acid capsule is a major virulence factor in the pathogenesis of Streptococcus pyogenes. It acts as an anti-phagocytic agent and adhesin to keratinocytes. The expression of the capsule is primarily regulated at the transcriptional level by the two-component regulatory system CovRS, in which CovR acts as a transcriptional repressor. The covRS genes are frequently mutated in many invasive strains, and a subset of the invasive CovRS mutants does not produce a detectable level of the capsule at 37 °C, but produces a significant amount of the capsule at sub-body temperatures. Here, we report that a prophage has a crucial role in this capsule thermoregulation. Passaging CovR-null strains showing capsule thermoregulation using a lab medium produced spontaneous mutants producing a significant amount of the capsule regardless of incubation temperature and this phenotypic change was caused by curing of a particular prophage. The lab strain HSC5 contains three prophages on the chromosome, and only ΦHSC5.3 was cured in all spontaneous mutants. This result indicates that the prophage ΦHSC5.3 plays a crucial role in capsule thermoregulation, most likely by repressing capsule production at 37 °C. PMID:27669311

  14. Resistance to macrolides in Streptococcus pyogenes in France in pediatric patients.

    PubMed

    Bingen, E; Fitoussi, F; Doit, C; Cohen, R; Tanna, A; George, R; Loukil, C; Brahimi, N; Le Thomas, I; Deforche, D

    2000-06-01

    A total of 1,500 recent throat isolates of Streptococcus pyogenes collected between 1996 and 1999 from children throughout France were tested for their susceptibility to erythromycin, azithromycin, josamycin, clindamycin, and streptogramin B. The erythromycin-resistant isolates were further studied for their genetic mechanism of resistance, by means of PCR. The clonality of these strains was also investigated by means of serotyping and ribotyping. In all, 6.2% of the strains were erythromycin resistant, and 3.4 and 2.8% expressed the constitutive MLS(B) and M resistance phenotypes and harbored the ermB and mefA genes, respectively; ermTR was recovered from one isolate which also harbored the ermB gene. Ten serotypes and 8 ribotypes were identified, but we identified 17 strains by combining serotyping with ribotyping. Among the eight ribotypes, the mefA gene was recovered from six clusters, one being predominant, while the ermB gene was recovered from four clusters, of which two were predominant.

  15. The ExPortal: an organelle dedicated to the biogenesis of secreted proteins in Streptococcus pyogenes.

    PubMed

    Rosch, Jason W; Caparon, Michael G

    2005-11-01

    The Gram-positive pathogen Streptococcus pyogenes secretes proteins through the ExPortal, a unique single microdomain of the cellular membrane specialized to contain the Sec translocons. It has been proposed that the ExPortal functions as an organelle to promote the biogenesis of secreted proteins by coordinating interactions between nascent unfolded secretory proteins and membrane-associated chaperones. In this study we provide evidence to support this model. It was found that HtrA (DegP), a surface anchored accessory factor required for maturation of the secreted SpeB cysteine protease, was localized exclusively to the ExPortal. Furthermore, the ATP synthase beta subunit was not localized to the ExPortal, suggesting that retention is likely restricted to a specific subset of exported proteins. Mutations that disrupted the anchoring, but not the protease activity, of HtrA, also altered the maturation kinetics of SpeB demonstrating that localization to the ExPortal was important for HtrA function. These data indicate that the ExPortal provides a mechanism by which Gram-positive bacteria can coordinate protein secretion and subsequent biogenesis in the absence of a specialized protein-folding compartment.

  16. Streptococcus pyogenes polymyxin B-resistant mutants display enhanced ExPortal integrity.

    PubMed

    Port, Gary C; Vega, Luis A; Nylander, Andrew B; Caparon, Michael G

    2014-07-01

    The ExPortal protein secretion organelle in Streptococcus pyogenes is an anionic phospholipid-containing membrane microdomain enriched in Sec translocons and postsecretion protein biogenesis factors. Polymyxin B binds to and disrupts ExPortal integrity, resulting in defective secretion of several toxins. To gain insight into factors that influence ExPortal organization, a genetic screen was conducted to select for spontaneous polymyxin B-resistant mutants displaying enhanced ExPortal integrity. Whole-genome resequencing of 25 resistant mutants revealed from one to four mutations per mutant genome clustered primarily within a core set of 10 gene groups. Construction of mutants with individual deletions or insertions demonstrated that 7 core genes confer resistance and enhanced ExPortal integrity through loss of function, while 3 were likely due to gain of function and/or combinatorial effects. Core resistance genes include a transcriptional regulator of lipid biosynthesis, several genes involved in nutrient acquisition, and a variety of genes involved in stress responses. Two members of the latter class also function as novel regulators of the secreted SpeB cysteine protease. Analysis of the most frequently isolated mutation, a single nucleotide deletion in a track of 9 consecutive adenine residues in pstS, encoding a component of a high-affinity Pi transporter, suggests that this sequence functions as a molecular switch to facilitate stress adaptation. Together, these data suggest the existence of a membrane stress response that promotes enhanced ExPortal integrity and resistance to cationic antimicrobial peptides.

  17. An Association Between Peptidoglycan Synthesis and Organization of the Streptococcus pyogenes ExPortal

    PubMed Central

    Vega, Luis Alberto; Port, Gary C.; Caparon, Michael G.

    2013-01-01

    ABSTRACT The ExPortal of Streptococcus pyogenes is a focal microdomain of the cytoplasmic membrane that clusters the translocons of the general secretory pathway with accessory factors to facilitate the maturation of secreted polypeptides. While it is known that the ExPortal is enriched in anionic lipids, the mechanisms that organize the ExPortal are poorly understood. In the present study, we examined the role of the cell wall in organizing and maintaining the ExPortal. Removal of the cell wall resulted in a loss of ExPortal focal integrity accompanied by the circumferential redistribution of ExPortal lipid and protein components. A similar loss occurred upon treatment with gallidermin, a nonpermeabilizing lantibiotic that targets the lipid II precursor of peptidoglycan synthesis, and this treatment disrupted the secretion of several ExPortal substrates. Furthermore, several enzymes involved in the membrane-associated steps of lipid II synthesis, including MraY and MurN, were found to localize to a single discrete focus in the membrane that was coincident with the focal location of the secretory translocons and the anionic lipid microdomain. These data suggest that the ExPortal is associated with the site of peptidoglycan precursor synthesis and that peptidoglycan biogenesis influences ExPortal organization. These data add to an emerging literature indicating that cell wall biogenesis, cell division, and protein secretion are spatially coorganized processes. PMID:24065630

  18. Cellular aspects of M protein and SfbI anchoring to Streptococcus pyogenes wall

    PubMed Central

    Raz, Assaf; Talay, Susanne; Fischetti, Vincent

    2012-01-01

    Summary Wall-anchored surface proteins are critical for the in vivo survival of Streptococcus pyogenes. Cues in the signal sequence direct the membrane translocation of surface proteins: M protein to the septum, and SfbI to the poles. Both proteins are subsequently anchored to the wall by the membrane bound enzyme sortase A. However, the cellular features of these pathways are not fully understood. Here we show that M protein and SfbI are anchored simultaneously throughout the cell cycle. M protein is rapidly anchored at the septum, and in part of the cell cycle, is anchored simultaneously at the mother and daughter septa. Conversely, SfbI accumulates gradually on peripheral peptidoglycan, resulting in a polar distribution. Sortase is not required for translocation of M protein or SfbI at their respective locations. Methicillin-induced unbalanced peptidoglycan synthesis diminishes surface M protein but not SfbI. Furthermore, overexpression of the division regulator DivIVA also diminishes surface M protein but increases SfbI. These results demonstrate a close connection between the regulation of cell division and protein anchoring. Better understanding of the spatial regulation of surface anchoring may lead to the identification of novel targets for the development of anti-infective agents, given the importance of surface molecules for pathogenesis. PMID:22512736

  19. Transcriptome Remodeling Contributes to Epidemic Disease Caused by the Human Pathogen Streptococcus pyogenes

    PubMed Central

    Beres, Stephen B.; Kachroo, Priyanka; Nasser, Waleed; Olsen, Randall J.; Zhu, Luchang; Flores, Anthony R.; de la Riva, Ivan; Paez-Mayorga, Jesus; Jimenez, Francisco E.; Cantu, Concepcion; Vuopio, Jaana; Jalava, Jari; Kristinsson, Karl G.; Gottfredsson, Magnus; Corander, Jukka; Fittipaldi, Nahuel; Di Luca, Maria Chiara; Petrelli, Dezemona; Vitali, Luca A.; Raiford, Annessa; Jenkins, Leslie

    2016-01-01

    ABSTRACT For over a century, a fundamental objective in infection biology research has been to understand the molecular processes contributing to the origin and perpetuation of epidemics. Divergent hypotheses have emerged concerning the extent to which environmental events or pathogen evolution dominates in these processes. Remarkably few studies bear on this important issue. Based on population pathogenomic analysis of 1,200 Streptococcus pyogenes type emm89 infection isolates, we report that a series of horizontal gene transfer events produced a new pathogenic genotype with increased ability to cause infection, leading to an epidemic wave of disease on at least two continents. In the aggregate, these and other genetic changes substantially remodeled the transcriptomes of the evolved progeny, causing extensive differential expression of virulence genes and altered pathogen-host interaction, including enhanced immune evasion. Our findings delineate the precise molecular genetic changes that occurred and enhance our understanding of the evolutionary processes that contribute to the emergence and persistence of epidemically successful pathogen clones. The data have significant implications for understanding bacterial epidemics and for translational research efforts to blunt their detrimental effects. PMID:27247229

  20. Multicenter study on invasive Streptococcus pyogenes infections in children in Argentina.

    PubMed

    Cancellara, Aldo D; Melonari, Pablo; Firpo, María V; Mónaco, Andrea; Ezcurra, Gustavo C; Ruizf, Lía; Aletti, Alicia M; Gregorio, Gabriela; Gaiano, Alejandra; Aird, Alberto; Bellone, Luciana; Calvari, Miriam; Torregrosa, Carolina; Morinigo, Sandra; Vozzan, María L; Tonetto, Ivana; Flynn, Luis P; Bidone, Nancy M; Russ, Carlota; Ellis, Alejandro

    2016-06-01

    Invasive Streptococcus pyogenes infections (ISpIs) cause a high morbidity and mortality, even at present; however, at a regional level there are few publications on this subject in the field of pediatrics. To describe the prevalence, predisposing factors and clinical characteristics of children hospitalized for ISpI, and analyze risk factors associated with bacteremia and lethality. Material and methods. Retrospective, descriptive study on ISpIs in children <18 years old hospitalized in the Pediatric Ward of 20 healthcare facilities across Argentina between 2010 and 2012. Assessed outcome measures: age, gender, early and late clinical sources of infection, prior chronic condition, predisposing factors, treatment and evolution. To describe the prevalence, predisposing factors and clinical characteristics of children hospitalized for ISpI, and analyze risk factors associated with bacteremia and lethality. Material and methods. Retrospective, descriptive study on ISpIs in children <18 years old hospitalized in the Pediatric Ward of 20 healthcare facilities across Argentina between 2010 and 2012. Assessed outcome measures: age, gender, early and late clinical sources of infection, prior chronic condition, predisposing factors, treatment and evolution. Most ISpIs were observed in patients without a prior chronic condition. The most common manifestation was, frequently with bacteremia, in the skin and soft tissue. A statistically significant association was observed between bacteremia and ≥2 early sources of infection and no surgery. Fatality rate, in association with streptococcal toxic shock syndrome and necrotizing fasciitis, was similar to that observed in other publications. Sociedad Argentina de Pediatría.

  1. Epidemiology of invasive Streptococcus pyogenes disease in Germany during 2003-2007.

    PubMed

    Imöhl, Matthias; Reinert, Ralf René; Ocklenburg, Christina; van der Linden, Mark

    2010-04-01

    A nationwide laboratory-based surveillance study of invasive Streptococcus pyogenes infections was conducted in Germany. Invasive isolates (n=586) were obtained between 2003 and 2007. Most isolates were obtained from blood (53.9%) or skin lesions (17.6%). The most common emm types were emm 1 (30.5%), emm 28 (18.3%) and emm 3 (9.6%). Overall, speA was positive in 45.9%, speC in 44.7% and ssa in 14.8% of isolates. SpeA was common in emm type 1 (100%) and emm type 3 (96.4%), whereas speC was often observed in emm type 28 (93.5%). The most frequent clinical manifestations included sepsis (40.1%), necrotizing fasciitis (20.8%) and streptococcal toxic shock syndrome (16.6%). All isolates were susceptible to penicillin G, cefotaxime and levofloxacin. Tetracycline shows the highest rate of resistant or intermediate isolates with 11.6%, followed by clarithromycin (5.5%) and clindamycin (1.2%). The most prominent trend is the reduction of tetracycline-nonsusceptible isolates from 18.6% in 2003 to 8.9% in 2007.

  2. Structural model for covalent adhesion of the Streptococcus pyogenes pilus through a thioester bond.

    PubMed

    Linke-Winnebeck, Christian; Paterson, Neil G; Young, Paul G; Middleditch, Martin J; Greenwood, David R; Witte, Gregor; Baker, Edward N

    2014-01-03

    The human pathogen Streptococcus pyogenes produces pili that are essential for adhesion to host surface receptors. Cpa, the adhesin at the pilus tip, was recently shown to have a thioester-containing domain. The thioester bond is believed to be important in adhesion, implying a mechanism of covalent attachment analogous to that used by human complement factors. Here, we have characterized a second active thioester-containing domain on Cpa, the N-terminal domain of Cpa (CpaN). Expression of CpaN in Escherichia coli gave covalently linked dimers. These were shown by x-ray crystallography and mass spectrometry to comprise two CpaN molecules cross-linked by the polyamine spermidine following reaction with the thioester bonds. This cross-linked CpaN dimer provides a model for the covalent attachment of Cpa to target receptors and thus the streptococcal pilus to host cells. Similar thioester domains were identified in cell wall proteins of other Gram-positive pathogens, suggesting that thioester domains are more widely used and provide a mechanism of adhesion by covalent bonding to target molecules on host cells that mimics that used by the human complement system to eliminate pathogens.

  3. Structural Model for Covalent Adhesion of the Streptococcus pyogenes Pilus through a Thioester Bond*

    PubMed Central

    Linke-Winnebeck, Christian; Paterson, Neil G.; Young, Paul G.; Middleditch, Martin J.; Greenwood, David R.; Witte, Gregor; Baker, Edward N.

    2014-01-01

    The human pathogen Streptococcus pyogenes produces pili that are essential for adhesion to host surface receptors. Cpa, the adhesin at the pilus tip, was recently shown to have a thioester-containing domain. The thioester bond is believed to be important in adhesion, implying a mechanism of covalent attachment analogous to that used by human complement factors. Here, we have characterized a second active thioester-containing domain on Cpa, the N-terminal domain of Cpa (CpaN). Expression of CpaN in Escherichia coli gave covalently linked dimers. These were shown by x-ray crystallography and mass spectrometry to comprise two CpaN molecules cross-linked by the polyamine spermidine following reaction with the thioester bonds. This cross-linked CpaN dimer provides a model for the covalent attachment of Cpa to target receptors and thus the streptococcal pilus to host cells. Similar thioester domains were identified in cell wall proteins of other Gram-positive pathogens, suggesting that thioester domains are more widely used and provide a mechanism of adhesion by covalent bonding to target molecules on host cells that mimics that used by the human complement system to eliminate pathogens. PMID:24220033

  4. Preparation and characterization of monomers to tetramers of a collagen-like domain from Streptococcus pyogenes.

    PubMed

    Peng, Yong Y; Stoichevska, Violet; Howell, Linda; Madsen, Soren; Werkmeister, Jerome A; Dumsday, Geoff J; Ramshaw, John A M

    2014-01-01

    The collagen like domain Scl2 from Streptococcus pyogenes has been proposed as a potential biomedical material. It is non-cytotoxic and non-immunogenic and can be prepared in good yield in fermentation. The Scl2 collagen domain is about a quarter of the length, 234 residues, of the main collagen type, mammalian type I collagen (1014 residues) that is currently used in biomedical devices. In the present study we have made constructs comprising 1 to 4 copies of the Scl2 collagen domain, plus these same constructs with a CysCys sequence at the C-terminal, analogous to that found in mammalian type III collagens. The yields of these constructs were examined from 2 L fermentation studies. The yields of both series declined with increasing size. Circular dichroism showed that the addition of further collagen domains did not lead to a change in the melting temperature compared to the monomer domain. Addition of the CysCys sequence led to a small additional stabilization of about 2-3°C for the monomer construct when the folding (V) domain was present.

  5. Phenotypic differentiation of Streptococcus pyogenes populations is induced by recombination-driven gene-specific sweeps

    PubMed Central

    Bao, Yun-Juan; Shapiro, B. Jesse; Lee, Shaun W.; Ploplis, Victoria A.; Castellino, Francis J.

    2016-01-01

    Genomic recombination plays an important role in driving adaptive evolution and population differentiation in bacteria. However, controversy exists as to the effects of recombination on population diversity and differentiation, i.e., recombination is frequent enough to sweep through the population at selected gene loci (gene-specific sweeps), or the recombination rate is low without interfering genome-wide selective sweeps. Observations supporting either view are sparse. Pathogenic bacteria causing infectious diseases are promising candidates to provide observations of recombination. However, phenotype-associated differentiations are usually vague among them due to diverse disease manifestations. Here we report a population genomic study of the group A Streptococcus pyogenes (GAS), a human pathogen with highly recombining genomes. By employing a genome-wide association study on single nucleotide polymorphisms (SNPs), we demonstrate a phenotypic differentiation of GAS, represented by separate clustering of two sublineages associated with niche-specific infections, i.e., skin infection and pharyngitis-induced acute rheumatic fever. By quantifying SNPs associated with the differentiation in a statistical and phylogenetic context, we propose that the phenotype-associated differentiation arose through recombination-driven gene-specific sweeps, rather than genome-wide sweeps. Our work provides a novel paradigm of phenotype-associated differentiation induced by gene-specific sweeps in a human pathogen and has implications for understanding of driving forces of bacterial evolution. PMID:27821851

  6. The Human Pathogen Streptococcus pyogenes Releases Lipoproteins as Lipoprotein-rich Membrane Vesicles.

    PubMed

    Biagini, Massimiliano; Garibaldi, Manuela; Aprea, Susanna; Pezzicoli, Alfredo; Doro, Francesco; Becherelli, Marco; Taddei, Anna Rita; Tani, Chiara; Tavarini, Simona; Mora, Marirosa; Teti, Giuseppe; D'Oro, Ugo; Nuti, Sandra; Soriani, Marco; Margarit, Immaculada; Rappuoli, Rino; Grandi, Guido; Norais, Nathalie

    2015-08-01

    Bacterial lipoproteins are attractive vaccine candidates because they represent a major class of cell surface-exposed proteins in many bacteria and are considered as potential pathogen-associated molecular patterns sensed by Toll-like receptors with built-in adjuvanticity. Although Gram-negative lipoproteins have been extensively characterized, little is known about Gram-positive lipoproteins. We isolated from Streptococcus pyogenes a large amount of lipoproteins organized in vesicles. These vesicles were obtained by weakening the bacterial cell wall with a sublethal concentration of penicillin. Lipid and proteomic analysis of the vesicles revealed that they were enriched in phosphatidylglycerol and almost exclusively composed of lipoproteins. In association with lipoproteins, a few hypothetical proteins, penicillin-binding proteins, and several members of the ExPortal, a membrane microdomain responsible for the maturation of secreted proteins, were identified. The typical lipidic moiety was apparently not necessary for lipoprotein insertion in the vesicle bilayer because they were also recovered from the isogenic diacylglyceryl transferase deletion mutant. The vesicles were not able to activate specific Toll-like receptor 2, indicating that lipoproteins organized in these vesicular structures do not act as pathogen-associated molecular patterns. In light of these findings, we propose to name these new structures Lipoprotein-rich Membrane Vesicles. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Inducer expulsion in Streptococcus pyogenes: properties and mechanism of the efflux reaction

    SciTech Connect

    Sutrina, S.L.; Reizer, J.; Saier, M.H Jr.

    1988-04-01

    Expulsion of preaccumulated methyl-..beta..-D-thiogalactoside-phosphate (TMG-P) from Streptococcus pyogenes is a two-step process comprising intracellular dephosphorylation of TMG-P followed by rapid efflux of the intracellularly formed free galactoside. The present study identifies the mechanism and the order and characterizes the temperature dependency of the efflux step. Unidirectional efflux of the intracellularly formed (/sup 14/C)TMG was only slightly affected when measured in the presence of unlabeled TMG (25 to 400 mM) in the extracellular medium. In contrast, pronounced inhibition of net efflux was observed in the presence of relatively low concentrations (1 to 16 mM) of extracellular (/sup 14/C)TMG. Since net efflux was nearly arrested when the external concentration of (/sup 14/C)TMG approached the intracellular concentration of this sugar, we propose that a facilitated diffusion mechanism is responsible for efflux and equilibration of TMG between the intracellular and extracellular milieus. The exit reaction was markedly dependent upon temperature, exhibited a high energy of activation (23 kcal (ca. 96 kJ) per mol), and followed first-order kinetics, indicating that the permease mediating this efflux was not saturated under the conditions of expulsion employed.

  8. The Human Pathogen Streptococcus pyogenes Releases Lipoproteins as Lipoprotein-rich Membrane Vesicles*

    PubMed Central

    Biagini, Massimiliano; Garibaldi, Manuela; Aprea, Susanna; Pezzicoli, Alfredo; Doro, Francesco; Becherelli, Marco; Taddei, Anna Rita; Tani, Chiara; Tavarini, Simona; Mora, Marirosa; Teti, Giuseppe; D'Oro, Ugo; Nuti, Sandra; Soriani, Marco; Margarit, Immaculada; Rappuoli, Rino; Grandi, Guido; Norais, Nathalie

    2015-01-01

    Bacterial lipoproteins are attractive vaccine candidates because they represent a major class of cell surface-exposed proteins in many bacteria and are considered as potential pathogen-associated molecular patterns sensed by Toll-like receptors with built-in adjuvanticity. Although Gram-negative lipoproteins have been extensively characterized, little is known about Gram-positive lipoproteins. We isolated from Streptococcus pyogenes a large amount of lipoproteins organized in vesicles. These vesicles were obtained by weakening the bacterial cell wall with a sublethal concentration of penicillin. Lipid and proteomic analysis of the vesicles revealed that they were enriched in phosphatidylglycerol and almost exclusively composed of lipoproteins. In association with lipoproteins, a few hypothetical proteins, penicillin-binding proteins, and several members of the ExPortal, a membrane microdomain responsible for the maturation of secreted proteins, were identified. The typical lipidic moiety was apparently not necessary for lipoprotein insertion in the vesicle bilayer because they were also recovered from the isogenic diacylglyceryl transferase deletion mutant. The vesicles were not able to activate specific Toll-like receptor 2, indicating that lipoproteins organized in these vesicular structures do not act as pathogen-associated molecular patterns. In light of these findings, we propose to name these new structures Lipoprotein-rich Membrane Vesicles. PMID:26018414

  9. Streptococcus pneumoniae and Streptococcus pyogenes resistant to macrolides but sensitive to clindamycin: a common resistance pattern mediated by an efflux system.

    PubMed Central

    Sutcliffe, J; Tait-Kamradt, A; Wondrack, L

    1996-01-01

    Macrolide-resistant Streptococcus pyogenes isolates from Finland, Australia, and the United Kingdom and, more recently, Streptococcus pneumoniae and S. pyogenes strains from the United States were shown to have an unusual resistance pattern to macrolides, lincosamides, and streptogramin B antibiotics. This pattern, referred to as M resistance, consists of susceptibility to clindamycin and streptogramin B antibiotics but resistance to 14- and 15-membered macrolides. An evaluation of the macrolide-lincosamide-streptogramin B resistance phenotypes among our streptococcal strains collected from 1993 to 1995 suggested that this unusual resistance pattern is not rare. Eighty-five percent (n = 66) of the S. pneumoniae and 75% (n = 28) of the S. pyogenes strains in our collection had an M phenotype. The mechanism of M resistance was not mediated by target modification, as isolated ribosomes from a pneumococcal strain bearing the M phenotype were fully sensitive to erythromycin. Further, the presence of an erm methylase was excluded with primers specific for an erm consensus sequence. However, results of studies that determined the uptake and incorporation of radiolabeled erythromycin into cells were consistent with the presence of a macrolide efflux determinant. The putative efflux determinant in streptococci seems to be distinct from the multicomponent macrolide efflux system in coagulase-negative staphylococci. The recognition of the prevalence of the M phenotype in streptococci has implications for sensitivity testing and may have an impact on the choice of antibiotic therapy in clinical practice. PMID:8843287

  10. Genome-scale reconstruction of the Streptococcus pyogenes M49 metabolic network reveals growth requirements and indicates potential drug targets.

    PubMed

    Levering, Jennifer; Fiedler, Tomas; Sieg, Antje; van Grinsven, Koen W A; Hering, Silvio; Veith, Nadine; Olivier, Brett G; Klett, Lara; Hugenholtz, Jeroen; Teusink, Bas; Kreikemeyer, Bernd; Kummer, Ursula

    2016-08-20

    Genome-scale metabolic models comprise stoichiometric relations between metabolites, as well as associations between genes and metabolic reactions and facilitate the analysis of metabolism. We computationally reconstructed the metabolic network of the lactic acid bacterium Streptococcus pyogenes M49. Initially, we based the reconstruction on genome annotations and already existing and curated metabolic networks of Bacillus subtilis, Escherichia coli, Lactobacillus plantarum and Lactococcus lactis. This initial draft was manually curated with the final reconstruction accounting for 480 genes associated with 576 reactions and 558 metabolites. In order to constrain the model further, we performed growth experiments of wild type and arcA deletion strains of S. pyogenes M49 in a chemically defined medium and calculated nutrient uptake and production fluxes. We additionally performed amino acid auxotrophy experiments to test the consistency of the model. The established genome-scale model can be used to understand the growth requirements of the human pathogen S. pyogenes and define optimal and suboptimal conditions, but also to describe differences and similarities between S. pyogenes and related lactic acid bacteria such as L. lactis in order to find strategies to reduce the growth of the pathogen and propose drug targets. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Analysis of the pathological lesions of the lung in a mouse model of cutaneous infection with Streptococcus pyogenes.

    PubMed

    Minami, Masaaki; Sobue, Sayaka; Ichihara, Masatoshi; Hasegawa, Tadao

    2012-02-01

    Invasive diseases such as toxic shock syndrome caused by Streptococcus pyogenes (S. pyogenes) are re-emerging infectious diseases. The mechanism of pathogenesis is not completely understood although the virulence of this organism has been analyzed using animal model systems, particularly using mice. The analysis of the progression of infection, however, is difficult. Computed tomography (CT) scanning is an extremely powerful technique that we applied to the mouse model of cutaneous infection with S. pyogenes. Two or three days after subcutaneous administration of bacteria, high density reticular areas were detected in the lung by CT. Histopathological examination of the lung was performed to examine the results of CT. Increased numbers of cytokeratin-positive epithelial cells, probably alveolar type II epithelial cells, were detected but no remarkable increase of inflammatory cell infiltrates was observed. Our results show that the pathological lesions of the lung in this model, wherein relatively few numbers of neutrophils were in the alveoli, are well correlated with the lung of a part of streptococcal toxic shock syndrome patients. Therefore, CT may be useful in assessing the progression of S. pyogenes infection, particularly in the pathological lesions of the lung in this model. © 2011 The Authors. Pathology International © 2011 Japanese Society of Pathology and Blackwell Publishing Asia Pty Ltd.

  12. Type I interferon production induced by Streptococcus pyogenes-derived nucleic acids is required for host protection.

    PubMed

    Gratz, Nina; Hartweger, Harald; Matt, Ulrich; Kratochvill, Franz; Janos, Marton; Sigel, Stefanie; Drobits, Barbara; Li, Xiao-Dong; Knapp, Sylvia; Kovarik, Pavel

    2011-05-01

    Streptococcus pyogenes is a Gram-positive human pathogen that is recognized by yet unknown pattern recognition receptors (PRRs). Engagement of these receptor molecules during infection with S. pyogenes, a largely extracellular bacterium with limited capacity for intracellular survival, causes innate immune cells to produce inflammatory mediators such as TNF, but also type I interferon (IFN). Here we show that signaling elicited by type I IFNs is required for successful defense of mice against lethal subcutaneous cellulitis caused by S. pyogenes. Type I IFN signaling was accompanied with reduced neutrophil recruitment to the site of infection. Mechanistic analysis revealed that macrophages and conventional dendritic cells (cDCs) employ different signaling pathways leading to IFN-beta production. Macrophages required IRF3, STING, TBK1 and partially MyD88, whereas in cDCs the IFN-beta production was fully dependent on IRF5 and MyD88. Furthermore, IFN-beta production by macrophages was dependent on the endosomal delivery of streptococcal DNA, while in cDCs streptococcal RNA was identified as the IFN-beta inducer. Despite a role of MyD88 in both cell types, the known IFN-inducing TLRs were individually not required for generation of the IFN-beta response. These results demonstrate that the innate immune system employs several strategies to efficiently recognize S. pyogenes, a pathogenic bacterium that succeeded in avoiding recognition by the standard arsenal of TLRs.

  13. Enzyme-linked immunosorbent assay for detection of type A streptococcal exotoxin: kinetics and regulation during growth of Streptococcus pyogenes.

    PubMed Central

    Houston, C W; Ferretti, J J

    1981-01-01

    We describe the detection and quantitation of type A streptococcal exotoxin (erythrogenic toxin, streptococcal pyrogenic exotoxin) by an enzyme-linked immunosorbent assay. This sensitive and specific technique detected microgram amounts of type A exotoxin and was useful for studying the kinetics and regulation of type A exotoxin production during the growth of Streptococcus pyogenes NY5. Maximum production of type A exotoxin was observed during the mid-log phase of growth, similar to the production of other streptococcal extracellular products. When S. pyogenes NY5 was grown at 42 degrees C, decreases in both growth and type A exotoxin production were observed. The results obtained when we studied the influence of nutrient additives and metal ions on the production of type A exotoxin led to the conclusion that none of these factors significantly affected type A exotoxin synthesis and that regulation was constitutive. Images PMID:7026447

  14. Single- and multistep resistance selection studies on the activity of retapamulin compared to other agents against Staphylococcus aureus and Streptococcus pyogenes.

    PubMed

    Kosowska-Shick, Klaudia; Clark, Catherine; Credito, Kim; McGhee, Pamela; Dewasse, Bonifacio; Bogdanovich, Tatiana; Appelbaum, Peter C

    2006-02-01

    Retapamulin had the lowest rate of spontaneous mutations by single-step passaging and the lowest parent and selected mutant MICs by multistep passaging among all drugs tested for all Staphylococcus aureus strains and three Streptococcus pyogenes strains which yielded resistant clones. Retapamulin has a low potential for resistance selection in S. pyogenes, with a slow and gradual propensity for resistance development in S. aureus.

  15. Single- and Multistep Resistance Selection Studies on the Activity of Retapamulin Compared to Other Agents against Staphylococcus aureus and Streptococcus pyogenes

    PubMed Central

    Kosowska-Shick, Klaudia; Clark, Catherine; Credito, Kim; McGhee, Pamela; Dewasse, Bonifacio; Bogdanovich, Tatiana; Appelbaum, Peter C.

    2006-01-01

    Retapamulin had the lowest rate of spontaneous mutations by single-step passaging and the lowest parent and selected mutant MICs by multistep passaging among all drugs tested for all Staphylococcus aureus strains and three Streptococcus pyogenes strains which yielded resistant clones. Retapamulin has a low potential for resistance selection in S. pyogenes, with a slow and gradual propensity for resistance development in S. aureus. PMID:16436741

  16. Tn5253 family integrative and conjugative elements carrying mef(I) and catQ determinants in Streptococcus pneumoniae and Streptococcus pyogenes.

    PubMed

    Mingoia, Marina; Morici, Eleonora; Morroni, Gianluca; Giovanetti, Eleonora; Del Grosso, Maria; Pantosti, Annalisa; Varaldo, Pietro E

    2014-10-01

    The linkage between the macrolide efflux gene mef(I) and the chloramphenicol inactivation gene catQ was first described in Streptococcus pneumoniae (strain Spn529), where the two genes are located in a module designated IQ element. Subsequently, two different defective IQ elements were detected in Streptococcus pyogenes (strains Spy029 and Spy005). The genetic elements carrying the three IQ elements were characterized, and all were found to be Tn5253 family integrative and conjugative elements (ICEs). The ICE from S. pneumoniae (ICESpn529IQ) was sequenced, whereas the ICEs from S. pyogenes (ICESpy029IQ and ICESpy005IQ, the first Tn5253-like ICEs reported in this species) were characterized by PCR mapping, partial sequencing, and restriction analysis. ICESpn529IQ and ICESpy029IQ were found to share the intSp 23FST81 integrase gene and an identical Tn916 fragment, whereas ICESpy005IQ has int5252 and lacks Tn916. All three ICEs were found to lack the linearized pC194 plasmid that is usually associated with Tn5253-like ICEs, and all displayed a single copy of a toxin-antitoxin operon that is typically contained in the direct repeats flanking the excisable pC194 region when this region is present. Two different insertion sites of the IQ elements were detected, one in ICESpn529IQ and ICESpy029IQ, and another in ICESpy005IQ. The chromosomal integration of the three ICEs was site specific, depending on the integrase (intSp 23FST81 or int5252). Only ICESpy005IQ was excised in circular form and transferred by conjugation. By transformation, mef(I) and catQ were cotransferred at a high frequency from S. pyogenes Spy005 and at very low frequencies from S. pneumoniae Spn529 and S. pyogenes Spy029.

  17. Differential compartmentalization of Streptococcus pyogenes virulence factors and host protein binding properties as a mechanism for host adaptation.

    PubMed

    Kilsgård, Ola; Karlsson, Christofer; Malmström, Erik; Malmström, Johan

    2016-11-01

    Streptococcus pyogenes is an important human pathogen responsible for substantial morbidity and mortality worldwide. Although S. pyogenes is a strictly human pathogen with no other known animal reservoir, several murine infection models exist to explore different aspects of the bacterial pathogenesis. Inoculating mice with wild-type S. pyogenes strains can result in the generation of new bacterial phenotypes that are hypervirulent compared to the original inoculum. In this study, we used a serial mass spectrometry based proteomics strategy to investigate if these hypervirulent strains have an altered distribution of virulence proteins across the intracellular, surface associated and secreted bacterial compartments and if any change in compartmentalization can alter the protein-protein interaction network between bacteria and host proteins. Quantitative analysis of the S. pyogenes surface and secreted proteomes revealed that animal passaged strains are associated with significantly higher amount of virulence factors on the bacterial surface and in the media. This altered virulence factor compartmentalization results in increased binding of several mouse plasma proteins to the bacterial surface, a trend that was consistent for mouse plasma from several different mouse strains. In general, both the wild-type strain and animal passaged strain were capable of binding high amounts of human plasma proteins. However, compared to the non-passaged strains, the animal passaged strains displayed an increased ability to bind mouse plasma proteins, in particular for M protein binders, indicating that the increased affinity for mouse blood plasma proteins is a consequence of host adaptation of this pathogen to a new host. In conclusion, plotting the total amount of virulence factors against the total amount of plasma proteins associated to the bacterial surface could clearly separate out animal passaged strains from wild type strains indicating a virulence model that could

  18. Bacterial Superantigens Promote Acute Nasopharyngeal Infection by Streptococcus pyogenes in a Human MHC Class II-Dependent Manner

    PubMed Central

    Kasper, Katherine J.; Zeppa, Joseph J.; Wakabayashi, Adrienne T.; Xu, Stacey X.; Mazzuca, Delfina M.; Welch, Ian; Baroja, Miren L.; Kotb, Malak; Cairns, Ewa; Cleary, P. Patrick; Haeryfar, S. M. Mansour; McCormick, John K.

    2014-01-01

    Establishing the genetic determinants of niche adaptation by microbial pathogens to specific hosts is important for the management and control of infectious disease. Streptococcus pyogenes is a globally prominent human-specific bacterial pathogen that secretes superantigens (SAgs) as ‘trademark’ virulence factors. SAgs function to force the activation of T lymphocytes through direct binding to lateral surfaces of T cell receptors and class II major histocompatibility complex (MHC-II) molecules. S. pyogenes invariably encodes multiple SAgs, often within putative mobile genetic elements, and although SAgs are documented virulence factors for diseases such as scarlet fever and the streptococcal toxic shock syndrome (STSS), how these exotoxins contribute to the fitness and evolution of S. pyogenes is unknown. Here we show that acute infection in the nasopharynx is dependent upon both bacterial SAgs and host MHC-II molecules. S. pyogenes was rapidly cleared from the nasal cavity of wild-type C57BL/6 (B6) mice, whereas infection was enhanced up to ∼10,000-fold in B6 mice that express human MHC-II. This phenotype required the SpeA superantigen, and vaccination with an MHC –II binding mutant toxoid of SpeA dramatically inhibited infection. Our findings indicate that streptococcal SAgs are critical for the establishment of nasopharyngeal infection, thus providing an explanation as to why S. pyogenes produces these potent toxins. This work also highlights that SAg redundancy exists to avoid host anti-SAg humoral immune responses and to potentially overcome host MHC-II polymorphisms. PMID:24875883

  19. Intra- and Interspecies Signaling between Streptococcus salivarius and Streptococcus pyogenes Mediated by SalA and SalA1 Lantibiotic Peptides

    PubMed Central

    Upton, M.; Tagg, J. R.; Wescombe, P.; Jenkinson, H. F.

    2001-01-01

    Streptococcus salivarius 20P3 produces a 22-amino-acid residue lantibiotic, designated salivaricin A (SalA), that inhibits the growth of a range of streptococci, including all strains of Streptococcus pyogenes. Lantibiotic production is associated with the sal genetic locus comprising salA, the lantibiotic structural gene; salBCTX genes encoding peptide modification and export machinery proteins; and salYKR genes encoding a putative immunity protein and two-component sensor-regulator system. Insertional inactivation of salB in S. salivarius 20P3 resulted in abrogation of SalA peptide production, of immunity to SalA, and of salA transcription. Addition of exogenous SalA peptide to salB mutant cultures induced dose-dependent expression of salA mRNA (0.2 kb), demonstrating that SalA production was normally autoregulated. Inactivation of salR encoding the response regulator of the SalKR two-component system led to reduced production of, and immunity to, SalA. The sal genetic locus was also present in S. pyogenes SF370 (M type 1), but because of a deletion across the salBCT genes, the corresponding lantibiotic peptide, designated SalA1, was not produced. However, in S. pyogenes T11 (M type 4) the sal locus gene complement was apparently complete, and active SalA1 peptide was synthesized. Exogenously added SalA1 peptide from S. pyogenes T11 induced salA1 transcription in S. pyogenes SF370 and in an isogenic S. pyogenes T11 salB mutant and salA transcription in S. salivarius 20P3 salB. Thus, SalA and SalA1 are examples of streptococcal lantibiotics whose production is autoregulated. These peptides act as intra- and interspecies signaling molecules, modulating lantibiotic production and possibly influencing streptococcal population ecology in the oral cavity. PMID:11395456

  20. [Evaluation of the safety and efficacy of cefditoren pivoxil fine granules for pediatric use in pediatric patients with laryngopharyngitis and tonsillitis caused by Streptococcus pyogenes].

    PubMed

    Kawamata, Saori; Yamada, Hitoshi; Sato, Yoshikazu; Sasagawa, Yuji; Iwama, Yasuhiro; Matumoto, Masato

    2010-08-01

    Rheumatic fever and acute glomerulonephritis are known to occur secondary to infection with Streptococcus pyogenes, and early elimination of the Streptococcus pyogenes by treatment with an appropriate antibiotic is required. Treatment with penicillins for 10 days has been recommended for Streptococcus pyogenes infections, but cephems are also now being used, and cefditoren pivoxil (CDTR-PI) is listed as one of the recommended drugs in the Guidelines for the Management of Respiratory Infectious Disease in Children in Japan 2007. We therefore conducted this study in order to collect appropriate use information in the clinical setting of CDTR-PI to treat Streptococcus pyogenes infections. In this study, 790 patients were enrolled in 147 institutions. Of them, 734 and 718 patients were chosen for safety and efficacy analysis, respectively. There were 11 adverse drug reactions in 11 patients, and the incidence of adverse drug reactions was 1.50% (11/734 patients). The most common adverse drug reactions were diarrhea and hematuria, and there were 3 events of each, but a positive urinalysis after administration were only obserbed without the clinical symptoms. With the exception of the 3 patients in which the patient did not return to the hospital and the outcome is unknown, the patients either recovered from all of the adverse drug reactions or they were relieved. No serious adverse drug reactions were reported in this study. The response rate was 98.5% for laryngopharyngitis (457/464 patients) and 98.4% (250/254 patients) for tonsillitis. Examination of the response rates according to patient background showed that they were high, 95% or more, in every group. The Streptococcus pyogenes eradication rate was 94.6% for laryngopharyngitis (194/205 patients) and 92.4% (110/119 patients) for tonsillitis. In summary, CDTR-PI exhibited excellent safety and efficacy in laryngopharyngitis and tonsillitis caused by Streptococcus pyogenes, and CDTR-PI was reconfirmed as a useful

  1. Protective Mechanisms of Respiratory Tract Streptococci against Streptococcus pyogenes Biofilm Formation and Epithelial Cell Infection

    PubMed Central

    Fiedler, Tomas; Riani, Catur; Koczan, Dirk; Standar, Kerstin

    2013-01-01

    Streptococcus pyogenes (group A streptococci [GAS]) encounter many streptococcal species of the physiological microbial biome when entering the upper respiratory tract of humans, leading to the question how GAS interact with these bacteria in order to establish themselves at this anatomic site and initiate infection. Here we show that S. oralis and S. salivarius in direct contact assays inhibit growth of GAS in a strain-specific manner and that S. salivarius, most likely via bacteriocin secretion, also exerts this effect in transwell experiments. Utilizing scanning electron microscopy documentation, we identified the tested strains as potent biofilm producers except for GAS M49. In mixed-species biofilms, S. salivarius dominated the GAS strains, while S. oralis acted as initial colonizer, building the bottom layer in mixed biofilms and thereby allowing even GAS M49 to form substantial biofilms on top. With the exception of S. oralis, artificial saliva reduced single-species biofilms and allowed GAS to dominate in mixed biofilms, although the overall two-layer structure was unchanged. When covered by S. oralis and S. salivarius biofilms, epithelial cells were protected from GAS adherence, internalization, and cytotoxic effects. Apparently, these species can have probiotic effects. The use of Affymetrix array technology to assess HEp-2 cell transcription levels revealed modest changes after exposure to S. oralis and S. salivarius biofilms which could explain some of the protective effects against GAS attack. In summary, our study revealed a protection effect of respiratory tract bacteria against an important airway pathogen and allowed a first in vitro insight into local environmental processes after GAS enter the respiratory tract. PMID:23241973

  2. Growth Phase-Dependent Modulation of Rgg Binding Specificity in Streptococcus pyogenes

    PubMed Central

    Anbalagan, Srivishnupriya; Dmitriev, Alexander; McShan, W. Michael; Dunman, Paul M.

    2012-01-01

    Streptococcus pyogenes Rgg is a transcriptional regulator that interacts with the cofactor LacD.1 to control growth phase-dependent expression of genes, including speB, which encodes a secreted cysteine protease. LacD.1 is thought to interact with Rgg when glycolytic intermediates are abundant in a manner that prevents Rgg-mediated activation of speB expression via binding to the promoter region. When the intermediates diminish, LacD.1 dissociates from Rgg and binds to the speB promoter to activate expression. The purpose of this study was to determine if Rgg bound to chromatin during the exponential phase of growth and, if so, to identify the binding sites. Rgg bound to 62 chromosomal sites, as determined by chromatin immunoprecipitation coupled with DNA microarrays. Thirty-eight were within noncoding DNA, including sites upstream of the genes encoding the M protein (M49), serum opacity factor (SOF), fibronectin-binding protein (SfbX49), and a prophage-encoded superantigen, SpeH. Each of these sites contained a promoter that was regulated by Rgg, as determined with transcriptional fusion assays. Purified Rgg also bound to the promoter regions of emm49, sof, and sfbX49 in vitro. Results obtained with a lacD.1 mutant showed that both LacD.1 and Rgg were necessary for the repression of emm49, sof, sfbX49, and speH expression. Overall, the results indicated that the DNA binding specificity of Rgg is responsive to environmental changes in a LacD.1-dependent manner and that Rgg and LacD.1 directly control virulence gene expression in the exponential phase of growth. PMID:22636768

  3. Streptococcus pyogenes Polymyxin B-Resistant Mutants Display Enhanced ExPortal Integrity

    PubMed Central

    Port, Gary C.; Vega, Luis A.; Nylander, Andrew B.

    2014-01-01

    The ExPortal protein secretion organelle in Streptococcus pyogenes is an anionic phospholipid-containing membrane microdomain enriched in Sec translocons and postsecretion protein biogenesis factors. Polymyxin B binds to and disrupts ExPortal integrity, resulting in defective secretion of several toxins. To gain insight into factors that influence ExPortal organization, a genetic screen was conducted to select for spontaneous polymyxin B-resistant mutants displaying enhanced ExPortal integrity. Whole-genome resequencing of 25 resistant mutants revealed from one to four mutations per mutant genome clustered primarily within a core set of 10 gene groups. Construction of mutants with individual deletions or insertions demonstrated that 7 core genes confer resistance and enhanced ExPortal integrity through loss of function, while 3 were likely due to gain of function and/or combinatorial effects. Core resistance genes include a transcriptional regulator of lipid biosynthesis, several genes involved in nutrient acquisition, and a variety of genes involved in stress responses. Two members of the latter class also function as novel regulators of the secreted SpeB cysteine protease. Analysis of the most frequently isolated mutation, a single nucleotide deletion in a track of 9 consecutive adenine residues in pstS, encoding a component of a high-affinity Pi transporter, suggests that this sequence functions as a molecular switch to facilitate stress adaptation. Together, these data suggest the existence of a membrane stress response that promotes enhanced ExPortal integrity and resistance to cationic antimicrobial peptides. PMID:24794568

  4. The Crystal Structure of Streptococcus pyogenes Uridine Phosphorylase Reveals a Distinct Subfamily of Nucleoside Phosphorylases

    SciTech Connect

    Tran, Timothy H.; Christoffersen, S.; Allan, Paula W.; Parker, William B.; Piskur, Jure; Serra, I.; Terreni, M.; Ealick, Steven E.

    2011-09-20

    Uridine phosphorylase (UP), a key enzyme in the pyrimidine salvage pathway, catalyzes the reversible phosphorolysis of uridine or 2'-deoxyuridine to uracil and ribose 1-phosphate or 2'-deoxyribose 1-phosphate. This enzyme belongs to the nucleoside phosphorylase I superfamily whose members show diverse specificity for nucleoside substrates. Phylogenetic analysis shows Streptococcus pyogenes uridine phosphorylase (SpUP) is found in a distinct branch of the pyrimidine subfamily of nucleoside phosphorylases. To further characterize SpUP, we determined the crystal structure in complex with the products, ribose 1-phosphate and uracil, at 1.8 {angstrom} resolution. Like Escherichia coli UP (EcUP), the biological unit of SpUP is a hexamer with an ?/? monomeric fold. A novel feature of the active site is the presence of His169, which structurally aligns with Arg168 of the EcUP structure. A second active site residue, Lys162, is not present in previously determined UP structures and interacts with O2 of uracil. Biochemical studies of wild-type SpUP showed that its substrate specificity is similar to that of EcUP, while EcUP is {approx}7-fold more efficient than SpUP. Biochemical studies of SpUP mutants showed that mutations of His169 reduced activity, while mutation of Lys162 abolished all activity, suggesting that the negative charge in the transition state resides mostly on uracil O2. This is in contrast to EcUP for which transition state stabilization occurs mostly at O4.

  5. Throat Carriage Rate and Antimicrobial Resistance of Streptococcus pyogenes In Rural Children in Argentina.

    PubMed

    Delpech, Gastón; Sparo, Mónica; Baldaccini, Beatriz; Pourcel, Gisela; Lissarrague, Sabina; García Allende, Leonardo

    2017-03-01

    The aim of this study was to determine the prevalence of asymptomatic carriers of group A β-hemolytic streptococci (GAS) in children living in a rural community and to investigate the association between episodes of acute pharyngitis and carrier status. Throat swabs were collected from September to November 2013 among children 5-13 years of age from a rural community (Maria Ignacia-Vela, Argentina). The phenotypic characterization of isolates was performed by conventional tests. Antimicrobial susceptibility was assayed for penicillin, tetracycline, chloramphenicol, erythromycin, and clindamycin (disk diffusion). The minimum inhibitory concentration was determined for penicillin, cefotaxime, tetracycline, and erythromycin. The carriage of β-hemolytic streptococci was detected in 18.1% of participants, with Streptococcus pyogenes in 18 participants followed by S. dysgalactiae ssp. equisimilis in 5. The highest proportion of GAS was found in 8 to 10-year-old children. No significant association between the number of episodes of acute pharyngitis suffered in the last year and the carrier state was detected (p>0.05). Tetracycline resistance (55.5%) and macrolide-resistant phenotypes (11.1%) were observed. Resistance to penicillin, cefotaxime, or chloramphenicol was not expressed in any streptococcal isolate. The present study demonstrated significant throat carriage of GAS and the presence of group C streptococci (S. dysgalactiae ssp. equisimilis) in an Argentinian rural population. These results point out the need for continuous surveillance of GAS and non-GAS carriage as well as of antimicrobial resistance in highly susceptible populations, such as school-aged rural children. An extended surveillance program including school-aged children from different cities should be considered to estimate the prevalence of GAS carriage in Argentina.

  6. Adaptive evolution of the Streptococcus pyogenes regulatory aldolase LacD.1.

    PubMed

    Cusumano, Zachary; Caparon, Michael

    2013-03-01

    In the human-pathogenic bacterium Streptococcus pyogenes, the tagatose bisphosphate aldolase LacD.1 likely originated through a gene duplication event and was adapted to a role as a metabolic sensor for regulation of virulence gene transcription. Although LacD.1 retains enzymatic activity, its ancestral metabolic function resides in the LacD.2 aldolase, which is required for the catabolism of galactose. In this study, we compared these paralogous proteins to identify characteristics correlated with divergence and novel function. Surprisingly, despite the fact that these proteins have identical active sites and 82% similarity in amino acid sequence, LacD.1 was less efficient at cleaving both fructose and tagatose bisphosphates. Analysis of kinetic properties revealed that LacD.1's adaptation was associated with a decrease in k(cat) and an increase in K(m). Construction and analysis of enzyme chimeras indicated that non-active-site residues previously associated with the variable activities of human aldolase isoenzymes modulated LacD.1's affinity for substrate. Mutant LacD.1 proteins engineered to have LacD.2-like levels of enzymatic efficiency lost the ability to function as regulators, suggesting that an alteration in efficiency was required for adaptation. In competition under growth conditions that mimic a deep-tissue environment, LacD.1 conferred a significant gain in fitness that was associated with its regulatory activity. Taken together, these data suggest that LacD.1's adaptation represents a form of neofunctionalization in which duplication facilitated the gain of regulatory function important for growth in tissue and pathogenesis.

  7. Adaptive Evolution of the Streptococcus pyogenes Regulatory Aldolase LacD.1

    PubMed Central

    Cusumano, Zachary

    2013-01-01

    In the human-pathogenic bacterium Streptococcus pyogenes, the tagatose bisphosphate aldolase LacD.1 likely originated through a gene duplication event and was adapted to a role as a metabolic sensor for regulation of virulence gene transcription. Although LacD.1 retains enzymatic activity, its ancestral metabolic function resides in the LacD.2 aldolase, which is required for the catabolism of galactose. In this study, we compared these paralogous proteins to identify characteristics correlated with divergence and novel function. Surprisingly, despite the fact that these proteins have identical active sites and 82% similarity in amino acid sequence, LacD.1 was less efficient at cleaving both fructose and tagatose bisphosphates. Analysis of kinetic properties revealed that LacD.1's adaptation was associated with a decrease in kcat and an increase in Km. Construction and analysis of enzyme chimeras indicated that non-active-site residues previously associated with the variable activities of human aldolase isoenzymes modulated LacD.1's affinity for substrate. Mutant LacD.1 proteins engineered to have LacD.2-like levels of enzymatic efficiency lost the ability to function as regulators, suggesting that an alteration in efficiency was required for adaptation. In competition under growth conditions that mimic a deep-tissue environment, LacD.1 conferred a significant gain in fitness that was associated with its regulatory activity. Taken together, these data suggest that LacD.1's adaptation represents a form of neofunctionalization in which duplication facilitated the gain of regulatory function important for growth in tissue and pathogenesis. PMID:23316044

  8. Cell wall polysaccharide biosynthesis by membrane fragments from Streptococcus pyogenes and stabilized L-form.

    PubMed

    Cohen, M; Panos, C

    1971-05-01

    The formation and composition of a cell wall rhamnose-containing polysaccharide by membrane fragments from Streptococcus pyogenes and its stabilized L-form were compared. Also, the effect of prior treatment on the ability of coccal whole-cell and membrane fragments to incorporate radioactivity from thymidine diphosphate-(14)C-rhamnose, and the results of subsequent attempts to remove labeled polysaccharide from such membranes are given. L-form membrane fragments were capable of only 10% uptake of (14)C-rhamnose from this nucleotide as compared with streptococcal membranes. However, once bound, both membrane fragments polymerized rhamnose to the same extent. These findings tend to negate the almost complete lack of polymeric rhamnose within the intact L-form as being due to the absence of membrane enzymes necessary for the transfer of rhamnose from a suitable precursor to membrane acceptor sites or enzymes responsible for rhamnose polymerization. Degradation of labeled rhamnose polysaccharide after isolation from coccal membranes by mild acid hydrolysis showed muramic acid and glucosamine to be attached. This same polysaccharide from L-form membrane fragments was devoid of amino sugars. These data suggest the possible involvement of amino sugars in the attachment of cell wall polymeric rhamnose to the streptococcal cytoplasmic membrane. The absence of attached amino sugars to rhamnose polysaccharide from L-form membrane fragments is discussed in terms of this organism's continued inability for new cell wall formation. The isolation, from streptococcal membrane fragments, of a polysaccharide containing rhamnose and amino sugars common to at least two different streptococcal cell wall-type polymers was demonstrated.

  9. Insidious manifestation of pyogenic liver abscess caused by Streptococcus intermedius and Micrococcus luteus: a case report.

    PubMed

    Ioannou, Antreas; Xenophontos, Eleni; Karatsi, Alexandra; Petrides, Christos; Kleridou, Maro; Zintilis, Chrysostomos

    2016-01-01

    Pyogenic liver abscesses are caused by various microorganisms and usually present with fever, abdominal pain, leukocytosis and liver enzyme abnormalities. This case presents the insidious manifestation of a pyogenic liver abscess in a 34-year-old immunocompetent male, where classical manifestations of a liver abscess were absent. The microorganisms cultured from the abscess belonged to oral cavity's and gastrointestinal tract's normal flora.

  10. Pleural empyema and streptococcal toxic shock syndrome due to Streptococcus pyogenes in a healthy Spanish traveler in Japan.

    PubMed

    Sakai, Tetsuya; Taniyama, Daisuke; Takahashi, Saeko; Nakamura, Morio; Takahashi, Takashi

    2017-01-01

    Group A Streptococcus (GAS, Streptococcus pyogenes) causes invasive infections including streptococcal toxic shock syndrome (STSS) and local infections. To our knowledge, this is the first report of a case of an invasive GAS infection with pneumonia and pleural empyema (PE) followed by STSS (disseminated intravascular coagulation [DIC] and acute renal insufficiency) in a healthy male adult. He received combined supportive therapies of PE drainage, anti-DIC agent, hemodialysis, and antimicrobials and eventually made a clinical recovery. GAS isolated from PE was found to have emm1/speA genes, suggestive of a pathogenic strain. Clinicians should be aware of the possibility of this disease entity (pneumonia, PE, and STSS) in healthy male adults as well as children and adult women.

  11. Trading Capsule for Increased Cytotoxin Production: Contribution to Virulence of a Newly Emerged Clade of emm89 Streptococcus pyogenes

    PubMed Central

    Zhu, Luchang; Olsen, Randall J.; Nasser, Waleed; de la Riva Morales, Ivan

    2015-01-01

    ABSTRACT Strains of emm89 Streptococcus pyogenes have become one of the major causes of invasive infections worldwide in the last 10 years. We recently sequenced the genome of 1,125 emm89 strains and identified three major phylogenetic groups, designated clade 1, clade 2, and the epidemic clade 3. Epidemic clade 3 strains, which now cause the great majority of infections, have two distinct genetic features compared to clade 1 and clade 2 strains. First, all clade 3 organisms have a variant 3 nga promoter region pattern, which is associated with increased production of secreted cytolytic toxins SPN (S. pyogenes NADase) and SLO (streptolysin O). Second, all clade 3 strains lack the hasABC locus mediating hyaluronic acid capsule synthesis, whereas this locus is intact in clade 1 and clade 2 strains. We constructed isogenic mutant strains that produce different levels of SPN and SLO toxins and capsule (none, low, or high). Here we report that emm89 strains with elevated toxin production are significantly more virulent than low-toxin producers. Importantly, we also show that capsule production is dispensable for virulence in strains that already produce high levels of SPN and SLO. Our results provide new understanding about the molecular mechanisms contributing to the rapid emergence and molecular pathogenesis of epidemic clade 3 emm89 S. pyogenes. PMID:26443457

  12. Streptococcus pyogenes Associated Post-traumatic Brodie’s Abscess of Cuboid: A Case Report and Review of Literature

    PubMed Central

    Amit, Priyadarshi; Maharajan, Karthikeyan; Varma, Bhaskar

    2015-01-01

    Introduction: Brodie’s abscess of cuboid bone is one of the rarest diagnosis in children which most often is hematogenous in origin. Although Streptococcus pyogenes has been uncommonly implicated as causative organism in other bones, it is not yet reported in the cuboid. Case Report: We report the case of 14-year-old boy who presented with a lytic lesion in the cuboid bone. It was preceded by a penetrating injury with a small iron nail. He was treated with simple curettage without the addition of bone graft. Frank pus present in the cavity in the cuboid bone grew S. pyogenes on bacterial culture. Symptoms resolved after 6 weeks of antibiotics, however, complete radiological healing was obtained after 9 months. Conclusion: Although very rare, S. pyogenes associated Brodie’s abscess should strongly be suspected in a posttraumatic lytic lesion in the cuboid bone and bone grafting is not always required for bone healing even in presence of large pus-filled cavity. PMID:27299080

  13. Inhibitory role of acyl homoserine lactones in hemolytic activity and viability of Streptococcus pyogenes M6 S165

    PubMed Central

    Saroj, Sunil D.; Holmer, Linda; Berengueras, Júlia M.; Jonsson, Ann-Beth

    2017-01-01

    Streptococcus pyogenes an adapted human pathogen asymptomatically colonizes the nasopharynx, among other polymicrobial communities. However, information on the events leading to the colonization and expression of virulence markers subject to interspecies and host-bacteria interactions are limited. The interference of acyl homoserine lactones (AHLs) with the hemolytic activity and viability of S. pyogenes M6 S165 was examined. AHLs, with fatty acid side chains ≥12 carbon atoms, inhibited hemolytic activity by downregulating the expression of the sag operon involved in the production of streptolysin S. Inhibitory AHLs upregulated the expression of transcriptional regulator LuxR. Electrophoretic mobility shift assays revealed the interaction of LuxR with the region upstream of sagA. AHL-mediated bactericidal activity observed at higher concentrations (mM range) was an energy-dependent process, constrained by the requirement of glucose and iron. Ferrichrome transporter FtsABCD facilitated transport of AHLs across the streptococcal membrane. The study demonstrates a previously unreported role for AHLs in S. pyogenes virulence. PMID:28303956

  14. Contribution of Secreted NADase and Streptolysin O to the Pathogenesis of Epidemic Serotype M1 Streptococcus pyogenes Infections.

    PubMed

    Zhu, Luchang; Olsen, Randall J; Lee, Jessica D; Porter, Adeline R; DeLeo, Frank R; Musser, James M

    2017-03-01

    Streptococcus pyogenes secretes many toxins that facilitate human colonization, invasion, and dissemination. NADase (SPN) and streptolysin O (SLO) are two toxins that play important roles in pathogenesis. We previously showed that increased production of SPN and SLO in epidemic serotype M1 and M89 S. pyogenes strains is associated with rapid intercontinental spread and enhanced virulence. The biological functions of SPN and SLO have been extensively studied using eukaryotic cell lines, but the relative contribution of each of these two toxins to pathogenesis of epidemic M1 or M89 strains remains unexplored. Herein, using a genetically representative epidemic M1 strain and a panel of isogenic mutant derivative strains, we evaluated the relative contributions of SPN and SLO toxins to virulence in mouse models of necrotizing myositis, bacteremia, and skin and soft tissue infection. We found that isogenic mutants lacking SPN, SLO, and both toxins are equally impaired in ability to cause necrotizing myositis. In addition, mutants lacking either SPN or SLO are significantly attenuated in the bacteremia and soft tissue infection models, and the mutant strain lacking production of both toxins is further attenuated. The mutant strain lacking both SPN and SLO production is severely attenuated in ability to resist killing by human polymorphonuclear leukocytes. We conclude that both SPN and SLO contribute significantly to S. pyogenes pathogenesis in these virulence assays. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  15. A cluster of ecthyma outbreaks caused by a single clone of invasive and highly infective Streptococcus pyogenes.

    PubMed

    Wasserzug, Oshri; Valinsky, Lea; Klement, Eyal; Bar-Zeev, Yael; Davidovitch, Nadav; Orr, Nadav; Korenman, Zina; Kayouf, Raid; Sela, Tamar; Ambar, Ruhama; Derazne, Estela; Dagan, Ron; Zarka, Salman

    2009-05-01

    Ecthyma is an invasive, ulcerated skin infection. Four ecthyma outbreaks occurred in different infantry units in the Israeli Defense Force from October 2004 through February 2005. Morbidity attack rates in the first 3 outbreaks were 89% (49 of 55 soldiers), 73% (32 of 44), and 82% (37 of 45). In the fourth outbreak, in which early intervention (antimicrobial treatment and improvement of hygiene) was applied, the attack rate was 25% (10 of 40 soldiers). In the first outbreak cluster, 4 soldiers experienced poststreptococcal glomerulonephritis, and 5 cases of systemic sequelae were recorded (1 case of severe septic shock, 3 cases of pneumonia, and 1 case of septic olecranon bursitis). Streptococcus pyogenes and Staphylococcus aureus were isolated from ecthyma sores, oropharynx, and anterior nares of affected and unaffected soldiers involved in all 4 outbreaks. Although the S. aureus isolates had different genomic profiles, >90% of S. pyogenes isolates were identified as belonging to a single clone, emm type 81, T type 8. Epidemiological investigation revealed that the hygiene levels of the soldiers and their living conditions were probably the most important cause for the difference in attack rates, wound severity, and systemic sequelae found between and within the units. Our study demonstrates the possible ramifications of the combination of a virulent and highly infective S. pyogenes strain and poor living conditions, and it emphasizes the importance of early intervention in such conditions.

  16. New macrolides active against Streptococcus pyogenes with inducible or constitutive type of macrolide-lincosamide-streptogramin B resistance.

    PubMed

    Fernandes, P B; Baker, W R; Freiberg, L A; Hardy, D J; McDonald, E J

    1989-01-01

    Macrolide-resistant bacteria can be classified as inducibly resistant or constitutively resistant. Inducibly resistant bacteria are resistant to 14-membered macrolides, such as erythromycin and clarithromycin (A-56268), but are susceptible to the 16-membered macrolides, such as tylosin and spiramycin, as well as to clindamycin. Constitutively resistant bacteria are resistant to macrolide-lincosamide-streptogramin B antibiotics. In this study, the MICs of several erythromycin and clarithromycin analogs against macrolide-susceptible and macrolide-resistant Streptococcus pyogenes strains were determined. Four 11,12-carbamate analogs of clarithromycin had lower MICs than erythromycin did against S. pyogenes with the inducible or constitutive type of macrolide-lincosamide-streptogramin B resistance. Five 11,12-carbonate analogs of erythromycin with modifications at the 4" position of cladinose had lower MICs than did erythromycin against S. pyogenes with the constitutive type of resistance, and one of these compounds, which had a naphthyl-glycyl substitution at the 4" position, had a lower MIC than erythromycin against both the inducibly resistant and constitutively resistant strains. Two analogs of erythromycin with a modification on the 4" position of cladinose had lower MICs than erythromycin did against the constitutively resistant organisms but not against the inducibly resistant organisms. Thus, 14-membered macrolides can be modified so as to confer a low MIC when tested in vitro.

  17. The effect of carbon dioxide on susceptibility testing of azithromycin, clarithromycin and roxithromycin against clinical isolates of Streptococcus pneumoniae and Streptococcus pyogenes by broth microdilution and the Etest: Artemis Project-first-phase study.

    PubMed

    Johnson, Jack; Bouchillon, Sam; Pontani, Dennis

    1999-06-01

    OBJECTIVE: To evaluate the effect of carbon dioxide on the susceptibility testing, using broth microdilution and the Etest (AB Biodisk, Solna, Sweden), of azithromycin, clarithromycin and roxithromycin against Streptococcus pneumoniae and Streptococcus pyogenes. METHODS: Fresh clinical isolates collected from 36 hospital laboratories in 12 countries were evaluated using the Etest in the presence of carbon dioxide. The isolates were retested under ambient conditions (absence of carbon dioxide) using broth microdilution and/or the Etest. RESULTS: Carbon dioxide falsely elevated azithromycin, clarithromycin and roxithromycin MIC90S for S. pneumoniae, determined by the Etest, approximately 12-fold. Also, the azithromycin MIC90 for S. pyogenes was increased fourfold; the effect was less marked for clarithromycin and roxithromycin. When isolates were retested in the absence of carbon dioxide, using the Etest or microdilution, susceptibilities to azithromycin were comparable to those to clarithromycin (S. pneumoniae, 93.4% versus 91.3%; S. pyogenes, 96.4% versus 95.8%). Both organisms were less susceptible to roxithromycin (S. pneumoniae, 71.3%; S. pyogenes, 85.7%). An internal standard control, consisting of 50 isolates each of S. pneumoniae, S. pyogenes and Haemophilus influenzae, confirmed that azithromycin susceptibility testing resulted in falsely elevated MICs. CONCLUSIONS: Carbon dioxide falsely elevated azithromycin MICs for S. pneumoniae and S. pyogenes, with an apparent reduction in susceptibility. When the in vitro activity of azithromycin and other macrolides against S. pneumoniae and S. pyogenes is being evaluated, awareness of the pH effect is essential.

  18. MALDI-TOF mass spectrometry as a tool for differentiation of invasive and noninvasive Streptococcus pyogenes isolates

    PubMed Central

    Moura, Hercules; Woolfitt, Adrian R; Carvalho, Maria G; Pavlopoulos, Antonis; Teixeira, Lucia M; Satten, Glen A; Barr, John R

    2008-01-01

    A novel mass spectral fingerprinting and proteomics approach using MALDI-TOF MS was applied to detect and identify protein biomarkers of group A Streptococcus (GAS) strains. Streptococcus pyogenes ATCC 700294 genome strain was compared with eight GAS clinical isolates to explore the ability of MALDI-TOF MS to differentiate isolates. Reference strains of other bacterial species were also analyzed and compared with the GAS isolates. MALDI preparations were optimized by varying solvents, matrices, plating techniques, and mass ranges for S. pyogenes ATCC 700294. Spectral variability was tested. A subset of common, characteristic, and reproducible biomarkers in the range of 2000–14 000 Da were detected, and they appeared to be independent of the culture media. Statistical analysis confirmed method reproducibility. Random Forest analysis of all selected GAS isolates revealed differences among most of them, and summed spectra were used for hierarchical cluster analysis. Specific biomarkers were found for each strain, and invasive GAS isolates could be differentiated. GAS isolates from cases of necrotizing fasciitis were clustered together and were distinct from isolates associated with noninvasive infections, despite their sharing the same emm type. Almost 30% of the biomarkers detected were tentatively identified as ribosomal proteins. PMID:18537829

  19. Integration of Genomic and Other Epidemiologic Data to Investigate and Control a Cross-Institutional Outbreak of Streptococcus pyogenes

    PubMed Central

    Chalker, Victoria J.; Smith, Alyson; Al-Shahib, Ali; Botchway, Stella; Macdonald, Emily; Daniel, Roger; Phillips, Sarah; Platt, Steven; Doumith, Michel; Tewolde, Rediat; Coelho, Juliana; Jolley, Keith A.; Underwood, Anthony

    2016-01-01

    Single-strain outbreaks of Streptococcus pyogenes infections are common and often go undetected. In 2013, two clusters of invasive group A Streptococcus (iGAS) infection were identified in independent but closely located care homes in Oxfordshire, United Kingdom. Investigation included visits to each home, chart review, staff survey, microbiologic sampling, and genome sequencing. S. pyogenes emm type 1.0, the most common circulating type nationally, was identified from all cases yielding GAS isolates. A tailored whole-genome reference population comprising epidemiologically relevant contemporaneous isolates and published isolates was assembled. Data were analyzed independently using whole-genome multilocus sequencing and single-nucleotide polymorphism analyses. Six isolates from staff and residents of the homes formed a single cluster that was separated from the reference population by both analytical approaches. No further cases occurred after mass chemoprophylaxis and enhanced infection control. Our findings demonstrate the ability of 2 independent analytical approaches to enable robust conclusions from nonstandardized whole-genome analysis to support public health practice. PMID:27192043

  20. High Prevalence of Macrolide-resistance and Molecular Characterization of Streptococcus pyogenes Isolates Circulating in China from 2009 to 2016.

    PubMed

    Lu, Binghuai; Fang, Yujie; Fan, Yanyan; Chen, Xingchun; Wang, Junrui; Zeng, Ji; Li, Yi; Zhang, Zhijun; Huang, Lei; Li, Hongxia; Li, Dong; Zhu, Fengxia; Cui, Yanchao; Wang, Duochun

    2017-01-01

    Streptococcus pyogenes, or group A Streptococcus, is a pathogen responsible for a wide range of clinical manifestations, from mild skin and soft tissue infections and pharyngitis to severe diseases. Its epidemiological characteristics should be comprehensively under surveillance for regulating the national prevention and treatment practice. Herein, a total of 140 S. pyogenes, including 38 invasive and 102 noninvasive isolates, were collected from infected patients in 10 tertiary general hospitals from 7 cities/provinces in China during the years 2009-2016. All strains were characterized by classical and molecular techniques for its emm types/subtypes, virulent factors and antibiotic resistance profiling. Of 140 isolates, 15 distinct emm types and 31 subtypes were detected, dominated by emm12 (60 isolates, 42.9%), emm1(43, 30.7%), and emm89 (10, 7.1%), and 8 new emm variant subtypes were identified. All strains, invasive or not, harbored the superantigenic genes, speB and slo. The other virulence genes, smeZ, speF, and speC accounted for 96.4, 91.4, and 87.1% of collected isolates, respectively. Further multilocus sequence typing (MLST) placed all strains into 22 individual sequence types (STs), including 4 newly-identified STs (11, 7.9%). All isolates were phenotypically susceptible to penicillin, ampicillin, cefotaxime, and vancomycin, whereas 131(93.5%), 132(94.2%), and 121(86.4%) were resistant to erythromycin, clindamycin, and tetracycline, respectively. Our study highlights high genotypic diversity and high prevalence of macrolide resistance of S. pyogenes among clinical isolates circulating in China.

  1. In vitro activity of midecamycin diacetate, a 16-membered macrolide, against Streptococcus pyogenes isolated in France, 1995-1999.

    PubMed

    Schlegel, L; Merad, B; Rostane, H; Broc, V; Bouvet, A

    2001-07-01

    To compare the in vitro activity of midecamycin diacetate to that of five other macrolides (erythromycin, clarithromycin, roxithromycin, azithromycin, and josamycin) and of clindamycin against 146 clinical isolates of Streptococcus pyogenes, with regard to three different phenotypes of erythromycin resistance. Susceptibility pattern and resistance phenotype were determined by disk diffusion method and double disk test. Minimal inhibitory concentrations of antibiotics were obtained by the agar dilution method and evaluated according to the recommendations of the 'Comité de l'Antibiogramme de la Société Française de Microbiologie' (CA-SFM). The major determinants of erythromycin resistance in S. pyogenes (ermB, ermTR and mefA genes) were investigated by specific amplification protocols. Most of the isolates of S. pyogenes collected during 1995-99 were susceptible to midecamycin (93.8%), erythromycin (90.4%), clarithromycin (93.2%), roxithromycin (91.8%), azithromycin (88.4%), josamycin (94.5%), and clindamycin (94.5%). According to the CA-SFM criteria, 132 of the 146 isolates studied were susceptible to erythromycin (MICs < or = 1 mg/L), four were intermediate (MICs 2-4 mg/L), and 10 were resistant (MICs > 4 mg/L). Only nine isolates were midecamycin resistant (MICs > 4 mg/L), and the others were susceptible. The increased activity of midecamycin (MIC90 < or = 0.06 mg/L), as compared to erythromycin (MIC90 = 0.5 mg/L) and to other 14- or 15-membered macrolides, was related to the absence of the ermB determinant in seven isolates which displayed an efflux phenotype (five isolates) or an inducible resistance phenotype due to an ermTR determinant (two isolates). Midecamycin diacetate is active against most S. pyogenes strains isolated in France and may represent an attractive alternative to the treatment of streptococcal infections due to resistant isolates with efflux of erythromycin.

  2. Structure and Activity of Streptococcus pyogenes SipA: A Signal Peptidase-Like Protein Essential for Pilus Polymerisation

    PubMed Central

    Young, Paul G.; Proft, Thomas; Harris, Paul W. R.; Brimble, Margaret A.; Baker, Edward N.

    2014-01-01

    The pili expressed on the surface of the human pathogen Streptococcus pyogenes play an important role in host cell attachment, colonisation and pathogenesis. These pili are built from two or three components, an adhesin subunit at the tip, a major pilin that forms a polymeric shaft, and a basal pilin that is attached to the cell wall. Assembly is carried out by specific sortase (cysteine transpeptidase) enzyme. These components are encoded in a small gene cluster within the S. pyogenes genome, often together with another protein, SipA, whose function is unknown. We show through functional assays, carried out by expressing the S. pyogenes pilus components in Lactococcus lactis, SipA from the clinically important M1T1 strain is essential for pilus assembly, and that SipA function is likely to be conserved in all S. pyogenes. From the crystal structure of SipA we confirm that SipA belongs to the family of bacterial signal peptidases (SPases), which process the signal-peptides of secreted proteins. In contrast to a previous arm-swapped SipA dimer, this present structure shows that its principal domain closely resembles the catalytic domain of SPases and has a very similar peptide-binding cleft, but it lacks the catalytic Ser and Lys residues characteristic of SPases. In SipA these are replaced by Asp and Gly residues, which play no part in activity. We propose that SipA functions by binding a key component at the bacterial cell surface, in a conformation that facilitates pilus assembly. PMID:24911348

  3. Fibronectin-binding protein of Streptococcus pyogenes: sequence of the binding domain involved in adherence of streptococci to epithelial cells.

    PubMed Central

    Talay, S R; Valentin-Weigand, P; Jerlström, P G; Timmis, K N; Chhatwal, G S

    1992-01-01

    The sequence of the fibronectin-binding domain of the fibronectin-binding protein of Streptococcus pyogenes (Sfb protein) was determined, and its role in streptococcal adherence was investigated by use of an Sfb fusion protein in adherence studies. A 1-kb DNA fragment coding for the binding domain of Sfb protein was cloned into the expression vector pEX31 to produce an Sfb fusion protein consisting of the N-terminal part of MS2 polymerase and a C-terminal fragment of the streptococcal protein. Induction of the vector promoter resulted in hyperexpression of fibronectin-binding fusion protein in the cytoplasm of the recombinant Escherichia coli cells. Sequence determination of the cloned 1-kb fragment revealed an in-frame reading frame for a 268-amino-acid peptide composed of a 37-amino-acid sequence which is completely repeated three times and incompletely repeated a fourth time. Cloning of one repeat into pEX31 resulted in expression of small fusion peptides that show fibronectin-binding activity, indicating that one repeat contains at least one binding domain. Each repeat exhibits two charged domains and shows high homology with the 38-amino-acid D3 repeat of the fibronectin-binding protein of Staphylococcus aureus. Sequence comparison with other streptococcal ligand-binding surface proteins, including M protein, failed to reveal significant homology, which suggests that Sfb protein represents a novel type of functional protein in S. pyogenes. The Sfb fusion protein isolated from the cytoplasm of recombinant cells was purified by fast protein liquid chromatography. It showed a strong competitive inhibition of fibronectin binding to S. pyogenes and of the adherence of bacteria to cultured epithelial cells. In contrast, purified streptococcal lipoteichoic acid showed only a weak inhibition of fibronectin binding and streptococcal adherence. These results demonstrate that Sfb protein is directly involved in the fibronectin-mediated adherence of S. pyogenes to

  4. Clinical evaluation of the oral probiotic Streptococcus salivarius K12 in the prevention of recurrent pharyngitis and/or tonsillitis caused by Streptococcus pyogenes in adults.

    PubMed

    Di Pierro, Francesco; Adami, Teresa; Rapacioli, Giuliana; Giardini, Nadia; Streitberger, Christian

    2013-03-01

    Streptococcus salivarius K12 has been shown to inhibit the growth of Streptococcus pyogenes due to bacteriocins release. Because of its ability to colonize the oral cavity, we have tested the strain K12 for its efficacy in preventing streptococcal pharyngitis and/or tonsillitis in adults. Forty adults with a diagnosis of recurrent oral streptococcal pharyngitis were enrolled in the study. Twenty of these subjects took for 90 days a tablet containing Streptococcus salivarius K12 (Bactoblis®). The other 20 subjects served as untreated controls. A 6-month follow-up was included to evaluate any persistent protective role. The 20 adults who completed the 90-day course of Bactoblis® showed a reduction in their episodes of streptococcal pharyngeal infection (about 80%). The 90 days treatment was also associated with an approximately 60% reduction in the incidence of reported pharyngitis in the 6-month period following use of the product. The product was well tolerated by the subjects with no treatment-related side effects or drop-outs reported. Prophylactic administration of Streptococcus salivarius K12 to adults having a history of recurrent oral streptococcal pathology reduced the number of episodes of streptococcal pharyngeal infections and/or tonsillitis.

  5. Structure and Kinetic Investigation of Streptococcus pyogenes Family GH38 α-Mannosidase

    PubMed Central

    Suits, Michael D. L.; Zhu, Yanping; Taylor, Edward J.; Walton, Julia; Zechel, David L.; Gilbert, Harry J.; Davies, Gideon J.

    2010-01-01

    Background The enzymatic hydrolysis of α−mannosides is catalyzed by glycoside hydrolases (GH), termed α−mannosidases. These enzymes are found in different GH sequence–based families. Considerable research has probed the role of higher eukaryotic “GH38” α−mannosides that play a key role in the modification and diversification of hybrid N-glycans; processes with strong cellular links to cancer and autoimmune disease. The most extensively studied of these enzymes is the Drosophila GH38 α−mannosidase II, which has been shown to be a retaining α−mannosidase that targets both α−1,3 and α−1,6 mannosyl linkages, an activity that enables the enzyme to process GlcNAc(Man)5(GlcNAc)2 hybrid N-glycans to GlcNAc(Man)3(GlcNAc)2. Far less well understood is the observation that many bacterial species, predominantly but not exclusively pathogens and symbionts, also possess putative GH38 α−mannosidases whose activity and specificity is unknown. Methodology/Principal Findings Here we show that the Streptococcus pyogenes (M1 GAS SF370) GH38 enzyme (Spy1604; hereafter SpGH38) is an α−mannosidase with specificity for α−1,3 mannosidic linkages. The 3D X-ray structure of SpGH38, obtained in native form at 1.9 Å resolution and in complex with the inhibitor swainsonine (Ki 18 µM) at 2.6 Å, reveals a canonical GH38 five-domain structure in which the catalytic “–1” subsite shows high similarity with the Drosophila enzyme, including the catalytic Zn2+ ion. In contrast, the “leaving group” subsites of SpGH38 display considerable differences to the higher eukaryotic GH38s; features that contribute to their apparent specificity. Conclusions/Significance Although the in vivo function of this streptococcal GH38 α−mannosidase remains unknown, it is shown to be an α−mannosidase active on N-glycans. SpGH38 lies on an operon that also contains the GH84 hexosaminidase (Spy1600) and an additional putative glycosidase. The activity of SpGH38, together

  6. Streptococcus pyogenes CAMP factor promotes bacterial adhesion and invasion in pharyngeal epithelial cells without serum via PI3K/Akt signaling pathway.

    PubMed

    Kurosawa, Mie; Oda, Masataka; Domon, Hisanori; Isono, Toshihito; Nakamura, Yuki; Saitoh, Issei; Hayasaki, Haruaki; Yamaguchi, Masaya; Kawabata, Shigetada; Terao, Yutaka

    2017-09-23

    Streptococcus pyogenes is a bacterium that causes systemic diseases, such as pharyngitis and toxic shock syndrome, via oral- or nasal-cavity infection. S. pyogenes produces various molecules known to function with serum components that lead to bacterial adhesion and invasion in human tissues. In this study, we identified a novel S. pyogenes adhesin/invasin. Our results revealed that CAMP factor promoted streptococcal adhesion and invasion in pharyngeal epithelial Detroit562 cells without serum. Recombinant CAMP factor initially localized on the membranes of cells and then became internalized in the cytosol following S. pyogenes infection. Additionally, CAMP factor phosphorylated phosphoinositide 3-kinase and serine-threonine kinase in the cells. ELISA results demonstrate that CAMP factor affected the amount of phosphorylated phosphoinositide 3-kinase and serine-threonine kinase in Detroit562 cells. Furthermore, CAMP factor did not reverse the effect of phosphoinositide 3-kinase knockdown by small interfering RNA in reducing the level of adhesion and invasion of S. pyogenes isogenic cfa-deficient mutant. These results suggested that S. pyogenes CAMP factor activated the phosphoinositide 3-kinase/serine-threonine kinase signaling pathway, promoting S. pyogenes invasion of Detroit562 cells without serum. Our findings suggested that CAMP factor played an important role on adhesion and invasion in pharyngeal epithelial cells. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  7. Role of putative virulence factors of Streptococcus pyogenes in mouse models of long-term throat colonization and pneumonia.

    PubMed Central

    Husmann, L K; Yung, D L; Hollingshead, S K; Scott, J R

    1997-01-01

    To investigate the role of putative virulence factors of Streptococcus pyogenes (group A streptococcus; GAS) in causing disease, we introduced specific mutations in GAS strain B514, a natural mouse pathogen, and tested the mutant strains in two models of infection. To study late stages of disease, we used our previously described mouse model (C3HeB/FeJ mice) in which pneumonia and systemic spread of the streptococcus follow intratracheal inoculation. To study the early stages of disease, we report here a model of long-term (at least 21 days) throat colonization following intranasal inoculation of C57BL/10SnJ mice. When the three emm family genes of GAS strain B514-Sm were deleted, the mutant showed no significant difference from the wild type in induction of long-term throat colonization or pneumonia. We inactivated the scpA gene, which encodes a complement C5a peptidase, by insertion of a nonreplicative plasmid and found no significant difference from the wild type in the incidence of throat colonization. However, there was a small but statistically significant decrease in the incidence of pneumonia caused by the scpA mutant. Finally, we demonstrated a very important effect of the hyaluronic acid capsule in both models. Following intranasal inoculation of mice with a mutant in which a nonreplicative plasmid was inserted into the hasA gene, which encodes hyaluronate synthase, we found that all bacteria recovered from the throats of the mice were encapsulated revertants. Following intratracheal inoculation with the hasA mutant, the incidence of pneumonia within 72 h was significantly reduced from that of the control strain (P = 0.006). These results indicate that the hyaluronic acid capsule of S. pyogenes B514 confers an important selective advantage for survival of the bacteria in the upper respiratory tract and is also an important determinant in induction of pneumonia in our model system. PMID:9119483

  8. In vitro induction and selection of fluoroquinolone-resistant mutants of Streptococcus pyogenes strains with multiple emm types.

    PubMed

    Billal, Dewan S; Fedorko, Daniel P; Yan, S Steve; Hotomi, Muneki; Fujihara, Keiji; Nelson, Nancy; Yamanaka, Noboru

    2007-01-01

    To perform a systematic analysis of point mutations in the quinolone resistance determining regions (QRDRs) of the DNA gyrase and topoisomerase genes of emm type 6 and other emm types of Streptococcus pyogenes strains after in vitro exposure to stepwise increasing concentrations of levofloxacin. Twelve parent strains of S. pyogenes, each with a different emm type, were chosen for stepwise exposure to increasing levels of levofloxacin followed by selection of resistant mutants. The QRDRs of gyrA, gyrB, parC and parE correlating to mutants with increased MICs were analysed for point mutations. Multiple mutants with significantly increased MICs were generated from each strain. The amino acid substitutions identified were consistent regardless of emm type and were similar to the mechanisms of resistance reported in clinical isolates of S. pyogenes. The number of induction/selection cycles required for the emergence of key point mutations in gyrA and parC was variable among strains. For each parent-mutant set, when MIC increased, serine-81 of gyrA and serine-79 of parC were the primary targets for amino acid substitutions. No point mutations were found in the QRDRs of gyrB and parE in any of the resistant mutants sequenced. Despite its intrinsic polymorphism in the QRDR of parC, emm type 6 is not more likely to develop high-level resistance to fluoroquinolones when compared with other emm types. All emm types seem equally inducible to high-level fluoroquinolone resistance.

  9. Exploration of fluoroquinolone resistance in Streptococcus pyogenes: comparative structure analysis of wild-type and mutant DNA gyrase.

    PubMed

    Shafreen, Raja Mohmed Beema; Selvaraj, Chandrabose; Singh, Sanjeev Kumar; Pandian, Shunmugiah Karutha

    2013-06-01

    Quinolone resistance-determining region is known to be the druggability site of the target protein that undergoes frequent mutation and thus renders quinolone resistance. In the present study, ligands were tested for their inhibitory activity against DNA gyrase of Streptococcus pyogenes involved in DNA replication. In silico mutational analysis on modelled gyrase A revealed that GLU85 had the most possible interactions with all the ligands used for the study. The amino acid residue GLU85 had also been predicted with an essential role of maintaining the three-dimensional structure of the protein. When introduced with a mutation (GLU 85 LYS) on this particular residue, it had readily denatured the whole α-helix (from 80 to 90 amino acids). This was confirmed through the molecular dynamics simulation and revealed that this single mutation can cause many functional and structural changes. Furthermore, LYS85 mutation has altered the original secondary structure of the protein, which in turn led to the steric hindrance during the ligand-receptor interaction. The results based on the G-score revealed that ligands have reduced interaction with the mutant protein. The semisynthetic fluoroquinolone 6d, which is an exception, forms a strong interaction with the mutant protein and was experimentally verified using the antimicrobial test. Hence, the present study unravels the fact that mutation at the drug binding site is the major cause for different level of resistance by the S. pyogenes when exposed against the varying concentrations of the fluoroquinolones. Furthermore, a comparative assessment of quinolone derivative with the older generation fluoroquinolones will be of great impact for S. pyogenes-related infections. Copyright © 2013 John Wiley & Sons, Ltd.

  10. The NADase-Negative Variant of the Streptococcus pyogenes Toxin NAD+ Glycohydrolase Induces JNK1-Mediated Programmed Cellular Necrosis

    PubMed Central

    Chandrasekaran, Sukantha

    2016-01-01

    ABSTRACT Virulence factors are often multifunctional and contribute to pathogenesis through synergistic mechanisms. For the human pathogen Streptococcus pyogenes, two factors that act synergistically are the S. pyogenes NAD+ glycohydrolase (SPN) and streptolysin O (SLO). Through distinct mechanisms, SLO forms pores in host cell membranes and translocates SPN into the host cell cytosol. Two natural variants of SPN exist, one that exhibits NADase activity and one that lacks this function, and both versions are translocated and act in concert with SLO to cause an accelerated death response in epithelial cells. While NADase+ SPN is known to trigger a metabolic form of necrosis through the depletion of NAD+, the mechanism by which NADase− SPN induces cell death was unknown. In the studies described here, we examined the pathway of NADase− cell death through analysis of activation patterns of mitogen-activated protein kinases (MAPKs). S. pyogenes infection resulted in activation of members of three MAPK subfamilies (p38, ERK, and JNK). However, only JNK was activated in an SLO-specific manner. NADase− SPN induced necrosis in HeLa epithelial cells associated with depolarization of mitochondrial membranes, activation of NF-κB, and the generation of reactive oxygen species. Remarkably, RNA interference (RNAi) silencing of JNK protected cells from NADase−-SPN-mediated necrosis, suggesting that NADase− SPN triggers a form of programmed necrosis dependent on JNK signaling. Taken together, these data demonstrate that SPN acts with SLO to elicit necrosis through two different mechanisms depending on its NADase activity, i.e., metabolic (NADase+) or programmed (NADase−), leading to distinct inflammatory profiles. PMID:26838722

  11. Distribution of mef(A)-containing genetic elements in erythromycin-resistant isolates of Streptococcus pyogenes from Italy.

    PubMed

    D'Ercole, S; Petrelli, D; Prenna, M; Zampaloni, C; Catania, M R; Ripa, S; Vitali, L A

    2005-11-01

    In total, 124 Streptococcus pyogenes isolates were obtained from throat cultures of different symptomatic patients. All isolates showed M-phenotype macrolide resistance and contained the macrolide efflux gene mef(A). The isolates were screened for the presence and insertion site of mef(A)-containing genetic elements. In 25.8% of the isolates, mef(A) was found to be carried by elements belonging to the Tn1207.3/Phi10394.4 family inserted in the comEC gene, while 74.2% contained chimeric elements with a different genetic structure and chromosomal location, probably associated with the recently described 60-kb tet(O)-mef(A) element.

  12. Correlation between genetic features of the mef(A)-msr(D) locus and erythromycin resistance in Streptococcus pyogenes.

    PubMed

    Vitali, Luca Agostino; Di Luca, Maria Chiara; Prenna, Manuela; Petrelli, Dezemona

    2016-01-01

    We investigated the correlation between the genetic variation within mef(A)-msr(D) determinants of efflux-mediated erythromycin resistance in Streptococcus pyogenes and the level of erythromycin resistance. Twenty-eight mef(A)-positive strains were selected according to erythromycin MIC (4-32 μg/mL), and their mef(A)-msr(D) regions were sequenced. Strains were classified according to the bacteriophage carrying mef(A)-msr(D). A new Φm46.1 genetic variant was found in 8 strains out of 28 and named VP_00501.1. Degree of allelic variation was higher in mef(A) than in msr(D). Hotspots for recombination were mapped within the locus that could have shaped the apparent mosaic structure of the region. There was a general correlation between mef(A)-msr(D) sequence and erythromycin resistance level. However, lysogenic conversion of susceptible strains by mef(A)-msr(D)-carrying Φm46.1 indicated that key determinants may not all reside within the mef(A)-msr(D) locus and that horizontal gene transfer could contribute to changes in the level of antibiotic resistance in S. pyogenes.

  13. Crystallization and preliminary X-ray crystallographic analysis of the tRNA-specific adenosine deaminase from Streptococcus pyogenes

    SciTech Connect

    Ku, Min-Je; Lee, Won-Ho; Nam, Ki-hyun; Rhee, Kyeong-hee; Lee, Ki-Seog; Kim, Eunice EunKyung; Yu, Myung-Hee; Hwang, Kwang Yeon

    2005-04-01

    The tRNA-specific adenosine deaminase from the pathogenic bacteria S. pyogenes has been overexpressed and crystallized. The tRNA-specific adenosine deaminase from the pathogenic bacteria Streptococcus pyogenes (spTAD) has been overexpressed in Escherichia coli and crystallized in the presence of Zn{sup 2+} ion at 295 K using ammonium sulfate as a precipitant. Flash-cooled crystals of spTAD diffracted to 2.0 Å using 30%(v/v) glycerol as a cryoprotectant. X-ray diffraction data have been collected to 2.0 Å using synchrotron radiation. The crystal belongs to the tetragonal space group P4{sub 2}2{sub 1}2, with unit-cell parameters a = b = 81.042, c = 81.270 Å. The asymmetric unit contains one subunit of spTAD, with a corresponding crystal volume per protein weight (V{sub M}) of 3.3 Å{sup 3} Da{sup −1} and a solvent content of 62.7%.

  14. The ScpC protease of Streptococcus pyogenes affects the outcome of sepsis in a murine model.

    PubMed

    Sjölinder, Hong; Lövkvist, Lena; Plant, Laura; Eriksson, Jens; Aro, Helena; Jones, Allison; Jonsson, Ann-Beth

    2008-09-01

    The ScpC protease of Streptococcus pyogenes degrades interleukin-8 (IL-8), a chemokine that mediates neutrophil transmigration and activation. The ability to degrade IL-8 differs dramatically among clinical isolates of S. pyogenes. Bacteria expressing ScpC overcome immune clearance by preventing the recruitment of neutrophils in soft tissue infection of mice. To study the role of ScpC in streptococcal sepsis, we generated an ScpC mutant that did not degrade IL-8 and thus failed to prevent the recruitment of immune cells as well as to cause disease after soft tissue infection. In a murine model of sepsis, challenge with the ScpC mutant resulted in more severe systemic disease with higher bacteremia levels and mortality than did challenge with the wild-type strain. As expected, the blood level of KC, the murine IL-8 homologue, increased in mice infected with the ScpC mutant. However, the elevated KC levels did not influence neutrophil numbers in blood, as it did in soft tissue, indicating that additional factors contributed to neutrophil transmigration in blood. In addition, the absence of ScpC increased tumor necrosis factor, IL-6, and C5a levels in blood, which contributed to disease severity. Thus, the ScpC mutant triggers high neutrophil infiltration but not lethal outcome after soft tissue infection, whereas intravenous infection leads to highly aggressive systemic disease.

  15. The YvqE two-component system controls biofilm formation and acid production in Streptococcus pyogenes.

    PubMed

    Isaka, Masanori; Tatsuno, Ichiro; Maeyama, Jun-Ichi; Matsui, Hideyuki; Zhang, Yan; Hasegawa, Tadao

    2016-07-01

    In Streptococcus pyogenes, proteins involved in determining virulence are controlled by stand-alone response regulators and by two-component regulatory systems. Previous studies reported that, compared to the parental strain, the yvqE sensor knockout strain showed significantly reduced growth and lower virulence. To determine the function of YvqE, we performed biofilm analysis and pH assays on yvqE mutants, and site-directed mutagenesis of YvqE. The yvqE deletion mutant showed a slower acid production rate, indicating that YvqE regulates acid production from sugar fermentation. The mutant strain, in which the Asp(26) residue in YvqE was replaced with Asn, affected biofilm formation, suggesting that this amino acid senses hydrogen ions produced by fermentative sugar metabolism. Signals received by YvqE were directly or indirectly responsible for inducing pilus expression. This study shows that at low environmental pH, biofilm formation in S. pyogenes is mediated by YvqE and suggests that regulation of pilus expression by environmental acidification could be directly under the control of YvqE. © 2016 APMIS. Published by John Wiley & Sons Ltd.

  16. Erythromycin resistance in italian isolates of Streptococcus pyogenes and correlations with pulsed-field gel electrophoresis analysis.

    PubMed

    Zampaloni, Claudia; Vitali, Luca A; Prenna, Manuela; Toscano, Maria A; Tempera, Gianna; Ripa, Sandro

    2002-01-01

    Erythromycin resistance among Streptococcus pyogenes strains has been reported in Italy at high rates during the last few years. A total of 152 erythromycin-resistant isolates of this species from southern Italian regions were characterized for the macrolide-resistance phenotype and screened by PCR for the corresponding genetic determinant. A close correlation was found between these phenotypic/genotypic data concerning macrolide resistance and results of Sma I macrorestriction fragment patterns (PFGE) analysis. In fact, the vast majority of the isolates assigned to individual PFGE classes mostly belonged to a single phenotype of macrolide resistance. All untypeable isolates belonged to the M phenotype. Twenty-two distinct PFGE types were recognized, of which 11 were recorded in only one isolate (one-strain type); about 50% of typeable isolates fell into five type clusters and 70% in seven. The increased erythromycin resistance among Italian isolates of S. pyogenes does not appear to be due to the spread of a single clone, but results indicate that the majority of group A streptococci examined are probably spread from a limited number of clones.

  17. [Invasive disease due to Streptococcus pyogenes in a patient with A H1N1 influenza infection. Report of one case].

    PubMed

    Guerrero S, Gonzalo; Marín S, Felipe

    2015-08-01

    Bacterial superinfection is a known complication among patients affected by viral respiratory tract infections. Streptococcus pyogenes, a major bacterial agent involved in acute tonsillopharyngitis, skin and soft tissue infections, was reported as a co-infecting microorganism during the 2009 A H1N1 influenza pandemic. We report a 65-year-old male patient who evolved with multifocal pneumonia and multiple organ failure with a fatal outcome. Influenza A H1N1 was detected by a polymerase chain reaction-based technique from a tracheal aspirate sample. S. pyogenes was identified by a rapid test from a nasopharyngeal sample and isolated afterwards from a positive blood culture.

  18. Activities of a New Fluoroketolide, HMR 3787, and Its (Des)-Fluor Derivative RU 64399 Compared to Those of Telithromycin, Erythromycin A, Azithromycin, Clarithromycin, and Clindamycin against Macrolide-Susceptible or -Resistant Streptococcus pneumoniae and S. pyogenes

    PubMed Central

    Nagai, Kensuke; Davies, Todd A.; Ednie, Lois M.; Bryskier, Andre; Palavecino, Elizabeth; Jacobs, Michael R.; Appelbaum, Peter C.

    2001-01-01

    Activities of HMR 3787 and RU 64399 were compared to those of three macrolides, telithromycin, and clindamycin against 175 Streptococcus pneumoniae isolates and 121 Streptococcus pyogenes isolates. HMR3787 and telithromycin were the most active compounds tested against pneumococci. Telithromycin and RU 64399 were equally active against macrolide-susceptible (MICs, 0.008 to 0.06 μg/ml) and -resistant S. pyogenes isolates, but HMR 3787 had lower MICs for ermB strains. PMID:11600391

  19. Citrulline Protects Streptococcus pyogenes from Acid Stress Using the Arginine Deiminase Pathway and the F1Fo-ATPase

    PubMed Central

    Cusumano, Zachary T.

    2015-01-01

    ABSTRACT A common stress encountered by both pathogenic and environmental bacteria is exposure to a low-pH environment, which can inhibit cell growth and lead to cell death. One major defense mechanism against this stress is the arginine deiminase (ADI) pathway, which catabolizes arginine to generate two ammonia molecules and one molecule of ATP. While this pathway typically relies on the utilization of arginine, citrulline has also been shown to enter into the pathway and contribute to protection against acid stress. In the pathogenic bacterium Streptococcus pyogenes, the utilization of citrulline has been demonstrated to contribute to pathogenesis in a murine model of soft tissue infection, although the mechanism underlying its role in infection is unknown. To gain insight into this question, we analyzed a panel of mutants defective in different steps in the ADI pathway to dissect how arginine and citrulline protect S. pyogenes in a low-pH environment. While protection provided by arginine utilization occurred through the buffering of the extracellular environment, citrulline catabolism protection was pH independent, requiring the generation of ATP via the ADI pathway and a functional F1Fo-ATP synthase. This work demonstrates that arginine and citrulline catabolism protect against acid stress through distinct mechanisms and have unique contributions to virulence during an infection. IMPORTANCE An important aspect of bacterial pathogenesis is the utilization of host-derived nutrients during an infection for growth and virulence. Previously published work from our lab identified a unique role for citrulline catabolism in Streptococcus pyogenes during a soft tissue infection. The present article probes the role of citrulline utilization during this infection and its contribution to protection against acid stress. This work reveals a unique and concerted action between the catabolism of citrulline and the F1Fo-ATPase that function together to provide protection for

  20. Antibiotic susceptibility in Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes in Pakistan: a review of results from the Survey of Antibiotic Resistance (SOAR) 2002-15.

    PubMed

    Zafar, A; Hasan, R; Nizamuddin, S; Mahmood, N; Mukhtar, S; Ali, F; Morrissey, I; Barker, K; Torumkuney, D

    2016-05-01

    To investigate changes in the antibiotic susceptibility of Streptococcus pneumoniae, Haemophilus influenzae and Streptococcus pyogenes from the Survey of Antibiotic Resistance (SOAR) in community-acquired respiratory tract infections (CA-RTIs) between 2002 and 2015 in Pakistan. This is a review based on previously published studies from 2002-03, 2004-06 and 2007-09 and also new data from 2014-15. Susceptibility was determined by Etest(®) or disc diffusion according to CLSI and pharmacokinetic/pharmacodynamic (PK/PD) breakpoints. A total of 706 isolates from CA-RTIs comprising 381 S. pneumoniae, 230 H. influenzae and 95 S. pyogenes were collected between 2002 and 2015 and tested against a range of antibiotics. Antibiotic resistance in S. pneumoniae rose steeply from 2002 to 2009, with isolates non-susceptible to penicillin and macrolides increasing from 10% to 34.1% and from 13%-14% to 29.7%, respectively. Susceptibility to amoxicillin/clavulanic acid (and by inference amoxicillin) remained between 99.4% and 100% from 2002 to 2015. Over the years, the prevalence of susceptibility to cefuroxime was 98%-100% among S. pneumoniae. Resistance in S. pneumoniae to some older antibiotics between 2007 and 2009 was high (86.8% for trimethoprim/sulfamethoxazole and 57.2% for tetracycline). Between 2002 and 2015, ampicillin resistance (β-lactamase-positive strains) among H. influenzae has remained low (between 2.6% and 3.2%) and almost unchanged over the years (H. influenzae was not tested during 2004-06). For S. pyogenes isolates, macrolide resistance reached 22%; however, susceptibility to penicillin, amoxicillin/clavulanic acid and cefuroxime remained stable at 100%. In S. pneumoniae from Pakistan, there has been a clear reduction in susceptibility to key antibiotics since 2002, but not to amoxicillin/clavulanic acid (amoxicillin) or cefuroxime. However, susceptibility in H. influenzae has remained stable. Local antibiotic susceptibility/resistance data are essential to

  1. Effects of the ERES pathogenicity region regulator Ralp3 on Streptococcus pyogenes serotype M49 virulence factor expression.

    PubMed

    Siemens, Nikolai; Fiedler, Tomas; Normann, Jana; Klein, Johannes; Münch, Richard; Patenge, Nadja; Kreikemeyer, Bernd

    2012-07-01

    Streptococcus pyogenes (group A streptococcus [GAS]) is a highly virulent Gram-positive bacterium. For successful infection, GAS expresses many virulence factors, which are clustered together with transcriptional regulators in distinct genomic regions. Ralp3 is a central regulator of the ERES region. In this study, we investigated the role of Ralp3 in GAS M49 pathogenesis. The inactivation of Ralp3 resulted in reduced attachment to and internalization into human keratinocytes. The Δralp3 mutant failed to survive in human blood and serum, and the hyaluronic acid capsule was slightly decreased. In addition, the mutant showed a lower binding capacity to human plasminogen, and the SpeB activity was significantly decreased. Complementation of the Δralp3 mutant restored the wild-type phenotype. The transcriptome and quantitative reverse transcription-PCR analysis of the serotype M49 GAS strain and its isogenic Δralp3 mutant identified 16 genes as upregulated, and 43 genes were found to be downregulated. Among the downregulated genes, there were open reading frames encoding proteins involved in metabolism (e.g., both lac operons and the fru operon), genes encoding lantibiotics (e.g., the putative salivaricin operon), and ORFs encoding virulence factors (such as the whole Mga core regulon and further genes under Mga control). In summary, the ERES region regulator Ralp3 is an important serotype-specific transcriptional regulator for virulence and metabolic control.

  2. SpyB, a small heme-binding protein, affects the composition of the cell wall in Streptococcus pyogenes

    DOE PAGES

    Edgar, Rebecca J.; Chen, Jing; Kant, Sashi; ...

    2016-10-13

    Streptococcus pyogenes (Group A Streptococcus or GAS) is a hemolytic human pathogen associated with a wide variety of infections ranging from minor skin and throat infections to life-threatening invasive diseases. The cell wall of GAS consists of peptidoglycan sacculus decorated with a carbohydrate comprising a polyrhamnose backbone with immunodominant N-acetylglucosamine side-chains. All GAS genomes contain the spyBA operon, which encodes a 35-amino-acid membrane protein SpyB, and a membrane-bound C3-like ADP-ribosyltransferase SpyA. In this study, we addressed the function of SpyB in GAS. Phenotypic analysis of a spyB deletion mutant revealed increased bacterial aggregation, and reduced sensitivity to β-lactams of themore » cephalosporin class and peptidoglycan hydrolase PlyC. Glycosyl composition analysis of cell wall isolated from the spyB mutant suggested an altered carbohydrate structure compared with the wild-type strain. Furthermore, we found that SpyB associates with heme and protoporphyrin IX. Heme binding induces SpyB dimerization, which involves disulfide bond formation between the subunits. Furthermore, our data suggest the possibility that SpyB activity is regulated by heme.« less

  3. Virulence factor regulation and regulatory networks in Streptococcus pyogenes and their impact on pathogen-host interactions.

    PubMed

    Kreikemeyer, Bernd; McIver, Kevin S; Podbielski, Andreas

    2003-05-01

    Streptococcus pyogenes (group A streptococcus, GAS) is a very important human pathogen with remarkable adaptation capabilities. Survival within the harsh host surroundings requires sensing potential on the bacterial side, which leads in particular to coordinately regulated virulence factor expression. GAS 'stand-alone' response regulators (RRs) and two-component signal transduction systems (TCSs) link the signals from the host environment with adaptive responses of the bacterial cell. Numerous putative regulatory systems emerged from GAS genome sequences. Only three RRs [Mga, RofA-like protein (RALP) and Rgg/RopB] and three TCSs (CsrRS/CovRS, FasBCAX and Ihk/Irr) have been studied in some detail with respect to their growth-phase-dependent activity and their influence on GAS-host cell interaction. In particular, the Mga-, RALP- and Rgg/RopB-regulated pathways display interconnected activities that appear to influence GAS colonization, persistence and spreading mechanisms, in a growth-phase-related fashion. Here, we have summarized our current knowledge about these RRs and TCSs to highlight the questions that should be addressed in future research on GAS pathogenicity.

  4. SpyB, a Small Heme-Binding Protein, Affects the Composition of the Cell Wall in Streptococcus pyogenes

    PubMed Central

    Edgar, Rebecca J.; Chen, Jing; Kant, Sashi; Rechkina, Elena; Rush, Jeffrey S.; Forsberg, Lennart S.; Jaehrig, Bernhard; Azadi, Parastoo; Tchesnokova, Veronika; Sokurenko, Evgeni V.; Zhu, Haining; Korotkov, Konstantin V.; Pancholi, Vijay; Korotkova, Natalia

    2016-01-01

    Streptococcus pyogenes (Group A Streptococcus or GAS) is a hemolytic human pathogen associated with a wide variety of infections ranging from minor skin and throat infections to life-threatening invasive diseases. The cell wall of GAS consists of peptidoglycan sacculus decorated with a carbohydrate comprising a polyrhamnose backbone with immunodominant N-acetylglucosamine side-chains. All GAS genomes contain the spyBA operon, which encodes a 35-amino-acid membrane protein SpyB, and a membrane-bound C3-like ADP-ribosyltransferase SpyA. In this study, we addressed the function of SpyB in GAS. Phenotypic analysis of a spyB deletion mutant revealed increased bacterial aggregation, and reduced sensitivity to β-lactams of the cephalosporin class and peptidoglycan hydrolase PlyC. Glycosyl composition analysis of cell wall isolated from the spyB mutant suggested an altered carbohydrate structure compared with the wild-type strain. Furthermore, we found that SpyB associates with heme and protoporphyrin IX. Heme binding induces SpyB dimerization, which involves disulfide bond formation between the subunits. Thus, our data suggest the possibility that SpyB activity is regulated by heme. PMID:27790410

  5. SpyB, a Small Heme-Binding Protein, Affects the Composition of the Cell Wall in Streptococcus pyogenes.

    PubMed

    Edgar, Rebecca J; Chen, Jing; Kant, Sashi; Rechkina, Elena; Rush, Jeffrey S; Forsberg, Lennart S; Jaehrig, Bernhard; Azadi, Parastoo; Tchesnokova, Veronika; Sokurenko, Evgeni V; Zhu, Haining; Korotkov, Konstantin V; Pancholi, Vijay; Korotkova, Natalia

    2016-01-01

    Streptococcus pyogenes (Group A Streptococcus or GAS) is a hemolytic human pathogen associated with a wide variety of infections ranging from minor skin and throat infections to life-threatening invasive diseases. The cell wall of GAS consists of peptidoglycan sacculus decorated with a carbohydrate comprising a polyrhamnose backbone with immunodominant N-acetylglucosamine side-chains. All GAS genomes contain the spyBA operon, which encodes a 35-amino-acid membrane protein SpyB, and a membrane-bound C3-like ADP-ribosyltransferase SpyA. In this study, we addressed the function of SpyB in GAS. Phenotypic analysis of a spyB deletion mutant revealed increased bacterial aggregation, and reduced sensitivity to β-lactams of the cephalosporin class and peptidoglycan hydrolase PlyC. Glycosyl composition analysis of cell wall isolated from the spyB mutant suggested an altered carbohydrate structure compared with the wild-type strain. Furthermore, we found that SpyB associates with heme and protoporphyrin IX. Heme binding induces SpyB dimerization, which involves disulfide bond formation between the subunits. Thus, our data suggest the possibility that SpyB activity is regulated by heme.

  6. A case of descending mediastinitis caused by Streptococcus pyogenes harboring genotype emm25 and sequence type 660.

    PubMed

    Ohya, Hiroaki; Mori, Nobuaki; Hayashi, Tetsuro; Minami, Shujiro; Higuchi, Akiko; Takahashi, Takashi

    2017-02-01

    Descending mediastinitis caused by Streptococcus pyogenes (group A streptococcus, GAS) is rare among cases of invasive GAS infection. In this report, we describe a case of a cervical abscess and secondary descending mediastinitis in a previously healthy 39-year-old Japanese man. The patient presented with a 2-week history of a sore throat, and subsequently developed an abscess and descending mediastinitis. We treated the cervical abscess using ampicillin/sulbactam and drainage, and GAS was subsequently isolated in two blood cultures from the patient's admission. Microbiological analyses revealed that the isolate harbored genotype emm25 and sequence type (ST) 660. This strain was susceptible to erythromycin (minimum inhibitory concentration [MIC]: ≤0.12 μg/mL), resistant to minocycline (MIC: >4 μg/mL), and possessed the tet(M) determinant. Although we have reviewed the literature regarding the clinical and microbiological characteristics of descending mediastinitis cause by GAS, little is known regarding epidemiological and clinical characteristics of emm25/ST660 GAS. Furthermore, to best of our knowledge, this is the first reported case of descending mediastinitis caused by emm25/ST660 GAS. Therefore, physicians should be aware of case with a cervical abscess and secondary descending mediastinitis caused by GAS infection, even if the patient is immunocompetent.

  7. Superoxide anions produced by Streptococcus pyogenes group A-stimulated keratinocytes are responsible for cellular necrosis and bacterial growth inhibition.

    PubMed

    Regnier, Elodie; Grange, Philippe A; Ollagnier, Guillaume; Crickx, Etienne; Elie, Laetitia; Chouzenoux, Sandrine; Weill, Bernard; Plainvert, Céline; Poyart, Claire; Batteux, Frédéric; Dupin, Nicolas

    2016-02-01

    Gram-positive Streptococcus pyogenes (group A Streptococcus or GAS) is a major skin pathogen and interacts with keratinocytes in cutaneous tissues. GAS can cause diverse suppurative and inflammatory infections, such as cellulitis, a common acute bacterial dermo-hypodermitis with a high morbidity. Bacterial isolation yields from the lesions are low despite the strong local inflammation observed, raising numerous questions about the pathogenesis of the infection. Using an in vitro model of GAS-infected keratinocytes, we show that the major ROS produced is the superoxide anion ([Formula: see text]), and that its production is time- and dose-dependent. Using specific modulators of ROS production, we show that [Formula: see text] is mainly synthesized by the cytoplasmic NADPH oxidase. Superoxide anion production leads to keratinocyte necrosis but incomplete inhibition of GAS growth, suggesting that GAS may be partially resistant to the oxidative burst. In conclusion, GAS-stimulated keratinocytes are able to develop an innate immune response based on the production of ROS. This local immune response limits GAS development and induces keratinocyte cell death, resulting in the skin lesions observed in patients with cellulitis. © The Author(s) 2015.

  8. [Multifocal invasive infection due to Streptococcus pyogenes in a 20 month-old infant: case report].

    PubMed

    Panetta, L; Manteau, C

    2014-11-01

    This case report describes a 20 months-old toddler with a multifocal S. pyogenes invasive infection cutaneous and articular, which outcome was favorable after implementation of a medical and surgical treatment. The antibiotic treatment was based on the association of amoxicillin and clindamycin which duration was guided by the clinical course and the secondary locations. A surgical management with joint irrigation lavage and drainage was necessary to induce a complete recovery. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  9. Necrotizing soft tissue infections caused by Streptococcus pyogenes and Streptococcus dysgalactiae subsp. equisimilis of groups C and G in western Norway.

    PubMed

    Bruun, T; Kittang, B R; de Hoog, B J; Aardal, S; Flaatten, H K; Langeland, N; Mylvaganam, H; Vindenes, H A; Skrede, S

    2013-12-01

    Streptococcus pyogenes (group A streptococcus, GAS) is a major cause of necrotizing soft tissue infection (NSTI). On rare occasions, other β-haemolytic streptococci may also cause NSTI, but the significance and nature of these infections has not been thoroughly investigated. In this study, clinical and molecular characteristics of NSTI caused by GAS and β-haemolytic Streptococcus dysgalactiae subsp. equisimilis of groups C and G (GCS/GGS) in western Norway during 2000-09 are presented. Clinical data were included retrospectively. The bacterial isolates were subsequently emm typed and screened for the presence of genes encoding streptococcal superantigens. Seventy cases were identified, corresponding to a mean annual incidence rate of 1.4 per 100 000. Sixty-one of the cases were associated with GAS, whereas GCS/GGS accounted for the remaining nine cases. The in-hospital case fatality rates of GAS and GCS/GGS disease were 11% and 33%, respectively. The GCS/GGS patients were older, had comorbidities more often and had anatomically more superficial disease than the GAS patients. High age and toxic shock syndrome were associated with mortality. The Laboratory Risk Indicator for Necrotizing Fasciitis laboratory score showed high values (≥6) in only 31 of 67 cases. Among the available 42 GAS isolates, the most predominant emm types were emm1, emm3 and emm4. The virulence gene profiles were strongly correlated to emm type. The number of superantigen genes was low in the four available GCS/GGS isolates. Our findings indicate a high frequency of streptococcal necrotizing fasciitis in our community. GCS/GGS infections contribute to the disease burden, but differ from GAS cases in frequency and predisposing factors. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  10. Complete Genome Sequence of emm28 Type Streptococcus pyogenes MEW123, a Streptomycin-Resistant Derivative of a Clinical Throat Isolate Suitable for Investigation of Pathogenesis

    PubMed Central

    Jacob, Kristin M.; Spilker, Theodore; LiPuma, John J.; Dawid, Suzanne R.

    2016-01-01

    We present here the complete genome sequence of Streptococcus pyogenes type emm28 strain MEW123, a streptomycin-resistant derivative of a pediatric throat isolate. The genome length is 1,878,699 bp, with 38.29% G+C% content. The genome sequence adds value to this virulent emm28 representative strain and will aid in the investigation of streptococcal pathogenesis. PMID:26988051

  11. Local activation of coagulation factor XIII reduces systemic complications and improves the survival of mice after Streptococcus pyogenes M1 skin infection.

    PubMed

    Deicke, Christin; Chakrakodi, Bhavya; Pils, Marina C; Dickneite, Gerhard; Johansson, Linda; Medina, Eva; Loof, Torsten G

    2016-11-01

    Coagulation is a mechanism for wound healing after injury. Several recent studies delineate an additional role of the intrinsic pathway of coagulation, also known as the contact system, in the early innate immune response against bacterial infections. In this study, we investigated the role of factor XIII (FXIII), which is activated upon coagulation induction, during Streptococcus pyogenes-mediated skin and soft tissue infections. FXIII has previously been shown to be responsible for the immobilization of bacteria within a fibrin network which may prevent systemic bacterial dissemination. In order to investigate if the FXIII-mediated entrapment of S. pyogenes also influences the disease outcome we used a murine S. pyogenes M1 skin and soft tissue infection model. Here, we demonstrate that a lack of FXIII leads to prolonged clotting times, increased signs of inflammation, and elevated bacterial dissemination. Moreover, FXIII-deficient mice show an impaired survival when compared with wildtype animals. Additionally, local reconstitution of FXIII-deficient mice with a human FXIII-concentrate (Fibrogammin(®)P) could reduce the systemic complications, suggesting a protective role for FXIII during early S. pyogenes skin infection. FXIII therefore might be a possible therapeutically application to support the early innate immune response during skin infections caused by S. pyogenes. Copyright © 2016 Elsevier GmbH. All rights reserved.

  12. Phim46.1, the main Streptococcus pyogenes element carrying mef(A) and tet(O) genes.

    PubMed

    Brenciani, Andrea; Bacciaglia, Alessandro; Vignaroli, Carla; Pugnaloni, Armanda; Varaldo, Pietro E; Giovanetti, Eleonora

    2010-01-01

    Phim46.1, the recognized representative of the most common variant of mobile, prophage-associated genetic elements carrying resistance genes mef(A) (which confers efflux-mediated erythromycin resistance) and tet(O) (which confers tetracycline resistance) in Streptococcus pyogenes, was fully characterized. Sequencing of the Phim46.1 genome (55,172 bp) demonstrated a modular organization typical of tailed bacteriophages. Electron microscopic analysis of mitomycin-induced Phim46.1 revealed phage particles with the distinctive icosahedral head and tail morphology of the Siphoviridae family. The chromosome integration site was within a 23S rRNA uracil methyltransferase gene. BLASTP analysis revealed that the proteins of Phim46.1 had high levels of amino acid sequence similarity to the amino acid sequences of proteins from other prophages, especially Phi10394.4 of S. pyogenes and lambdaSa04 of S. agalactiae. Phage DNA was present in the host cell both as a prophage and as free circular DNA. The lysogeny module appears to have been split due to the insertion of a segment containing tet(O) (from integrated conjugative element 2096-RD.2) and mef(A) (from a Tn1207.1-like transposon) into the unintegrated phage DNA. The phage attachment sequence lies in the region between tet(O) and mef(A) in the unintegrated form. Thus, whereas in this form tet(O) is approximately 5.5 kb upstream of mef(A), in the integrated form, tet(O), which lies close to the right end of the prophage, is approximately 46.3 kb downstream of mef(A), which lies close to the left end of the prophage.

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

    PubMed Central

    McNeilly, Celia; Cosh, Samantha; Vu, Therese; Nichols, Jemma; Henningham, Anna; Hofmann, Andreas; Fane, Anne; Smeesters, Pierre R.; Rush, Catherine M.; Hafner, Louise M.; Ketheesan, Natkuman; Sriprakash, Kadaba S.; McMillan, David J.

    2016-01-01

    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. PMID:27310707

  14. Emergence of Streptococcus pyogenes emm102 causing toxic shock syndrome in Southern Taiwan during 2005-2012.

    PubMed

    Lin, Jiun-Nong; Chang, Lin-Li; Lai, Chung-Hsu; Lin, Hsi-Hsun; Chen, Yen-Hsu

    2013-01-01

    Streptococcal toxic shock syndrome (STSS) is an uncommon but life-threatening disease caused by Streptococcus pyogenes. To understand the clinical and molecular characteristics of STSS, we analyzed clinical data and explored the emm types, superantigen genes, and pulsed-field gel electrophoresis of causative S. pyogenes isolates obtained between 2005 and 2012. In total, 53 patients with STSS were included in this study. The median age of the patients was 57 years (range: 9-83 years), and 81.1% were male. The most prevalent underlying disease was diabetes mellitus (45.3%). Skin and soft-tissue infection accounted for 86.8% of STSS. The overall mortality rate was 32.1%. Underlying diseases had no statistical impact on mortality. A total of 19 different emm types were identified. The most prevalent emm type was emm102 (18.9%), followed by emm11 (17%), emm1 (11.3%), emm87 (9.4%), and emm89 (7.5%). There was no statistically significant association between emm type and a fatal outcome. Among the superantigen genes, speB was the most frequently detected one (92.5%), followed by smeZ (90.6%), speG (81.1%), speC (39.6%), and speF (39.6%). The majority of emm102 strains were found to have speB, speC, speG, and smeZ. The presence of speG was negatively associated with a fatal outcome (P = 0.045). Our surveillance revealed the emergence of uncommon emm types, particularly emm102, causing STSS in southern Taiwan. Characterization of clinical, epidemiological, and molecular characteristics of STSS will improve our understanding of this life-threatening disease.

  15. Characterization of antimicrobial resistance in Streptococcus pyogenes isolates from the San Francisco Bay area of northern California.

    PubMed

    York, M K; Gibbs, L; Perdreau-Remington, F; Brooks, G F

    1999-06-01

    During 1994 and 1995, 157 isolates of Streptococcus pyogenes from patients with invasive disease were consecutively collected in the San Francisco Bay area to determine the frequency of antimicrobial resistance. Susceptibility testing was performed according to the guidelines of the National Committee for Clinical Laboratory Standards by the disk method and by broth microdilution. For comparison of susceptibility patterns, an additional 149 strains were randomly collected from patients with pharyngitis. For San Francisco County, 32% of the isolates from invasive-disease-related specimens but only 9% of the isolates from throat cultures from the same period were resistant to erythromycin (P = 0.0007). Alameda County and Contra Costa County had rates of resistance of pyogenes isolates

  16. Multiple Length Peptide-Pheromone Variants Produced by Streptococcus pyogenes Directly Bind Rgg Proteins to Confer Transcriptional Regulation*

    PubMed Central

    Aggarwal, Chaitanya; Jimenez, Juan Cristobal; Nanavati, Dhaval; Federle, Michael J.

    2014-01-01

    Streptococcus pyogenes, a human-restricted pathogen, accounts for substantial mortality related to infections worldwide. Recent studies indicate that streptococci produce and respond to several secreted peptide signaling molecules (pheromones), including those known as short hydrophobic peptides (SHPs), to regulate gene expression by a quorum-sensing mechanism. Upon transport into the bacterial cell, pheromones bind to and modulate activity of receptor proteins belonging to the Rgg family of transcription factors. Previously, we reported biofilm regulation by the Rgg2/3 quorum-sensing circuit in S. pyogenes. The aim of this study was to identify the composition of mature pheromones from cell-free culture supernatants that facilitate biofilm formation. Bioluminescent reporters were employed to detect active pheromones in culture supernatants fractionated by reverse-phase chromatography, and mass spectrometry was used to characterize their properties. Surprisingly, multiple SHPs that varied by length were detected. Synthetic peptides of each variant were tested individually using bioluminescence reporters and biofilm growth assays, and although activities differed widely among the group, peptides comprising the C-terminal eight amino acids of the full-length native peptide were most active. Direct Rgg/SHP interactions were determined using a fluorescence polarization assay that utilized FITC-labeled peptide ligands. Peptide receptor affinities were seen to be as low as 500 nm and their binding affinities directly correlated with observed bioactivity. Revelation of naturally produced pheromones along with determination of their affinity for cognate receptors are important steps forward in designing compounds whose purpose is positioned for future therapeutics aimed at treating infections through the interference of bacterial communication. PMID:24958729

  17. Multiple length peptide-pheromone variants produced by Streptococcus pyogenes directly bind Rgg proteins to confer transcriptional regulation.

    PubMed

    Aggarwal, Chaitanya; Jimenez, Juan Cristobal; Nanavati, Dhaval; Federle, Michael J

    2014-08-08

    Streptococcus pyogenes, a human-restricted pathogen, accounts for substantial mortality related to infections worldwide. Recent studies indicate that streptococci produce and respond to several secreted peptide signaling molecules (pheromones), including those known as short hydrophobic peptides (SHPs), to regulate gene expression by a quorum-sensing mechanism. Upon transport into the bacterial cell, pheromones bind to and modulate activity of receptor proteins belonging to the Rgg family of transcription factors. Previously, we reported biofilm regulation by the Rgg2/3 quorum-sensing circuit in S. pyogenes. The aim of this study was to identify the composition of mature pheromones from cell-free culture supernatants that facilitate biofilm formation. Bioluminescent reporters were employed to detect active pheromones in culture supernatants fractionated by reverse-phase chromatography, and mass spectrometry was used to characterize their properties. Surprisingly, multiple SHPs that varied by length were detected. Synthetic peptides of each variant were tested individually using bioluminescence reporters and biofilm growth assays, and although activities differed widely among the group, peptides comprising the C-terminal eight amino acids of the full-length native peptide were most active. Direct Rgg/SHP interactions were determined using a fluorescence polarization assay that utilized FITC-labeled peptide ligands. Peptide receptor affinities were seen to be as low as 500 nm and their binding affinities directly correlated with observed bioactivity. Revelation of naturally produced pheromones along with determination of their affinity for cognate receptors are important steps forward in designing compounds whose purpose is positioned for future therapeutics aimed at treating infections through the interference of bacterial communication. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. An Intrinsic Pattern of Reduced Susceptibility to Fluoroquinolones in Pediatric Isolates of Streptococcus pyogenes

    PubMed Central

    Yan, S. Steve; Schreckenberger, Paul C.; Zheng, Xiaotian; Nelson, Nancy A.; Harrington, Susan M.; Tjhio, Joyce; Fedorko, Daniel P.

    2008-01-01

    A total of 116 clinical isolates collected in 2003 from a tertiary pediatric hospital and a primary pediatric department in Chicago, Illinois were screened for reduced susceptibility to selected fluoroquinolones by disc diffusion. Correlation between reduced susceptibility and point mutations in the quinolone resistance-determining region of parC and gyrA genes were evaluated, and point mutations were compared with other reports of isolates derived from adult or mixed patient populations. 9% of isolates had reduced susceptibility to one or more of these fluoroquinolones by Etest: ciprofloxacin, levofloxacin, moxifloxacin. A single point mutation (Ser-79) in parC seemed responsible for the reduced susceptibility. Resistant S. pyogenes isolates were compared using M/emm type, RepPCR, and pulsed-field gel electrophoresis (PFGE). RepPCR provided no more separation of strains than M/emm typing and PFGE results with SgrA1 were more discriminatory than with SmaI. The majority of these isolates were M/emm type 6. PFGE analysis using SgrA1 demonstrated 2 different resistant strains among the M/emm type 6 isolates. The findings suggest that a population of S. pyogenes with an intrinsic reduced susceptibility to fluoroquinolones exists in pediatric clinical isolates. Monitoring of amino acid changes in both parC and gyrA will assist in the prediction of emergence of high level fluoroquinolone resistance. PMID:18554840

  19. A Highly Active and Negatively Charged Streptococcus pyogenes Lysin with a Rare d-Alanyl-l-Alanine Endopeptidase Activity Protects Mice against Streptococcal Bacteremia

    PubMed Central

    Lood, Rolf; Raz, Assaf; Molina, Henrik; Euler, Chad W.

    2014-01-01

    Bacteriophage endolysins have shown great efficacy in killing Gram-positive bacteria. PlyC, a group C streptococcal phage lysin, represents the most efficient lysin characterized to date, with a remarkably high specificity against different streptococcal species, including the important pathogen Streptococcus pyogenes. However, PlyC is a unique lysin, in terms of both its high activity and structure (two distinct subunits). We sought to discover and characterize a phage lysin active against S. pyogenes with an endolysin architecture distinct from that of PlyC to determine if it relies on the same mechanism of action as PlyC. In this study, we identified and characterized an endolysin, termed PlyPy (phage lysin from S. pyogenes), from a prophage infecting S. pyogenes. By in silico analysis, PlyPy was found to have a molecular mass of 27.8 kDa and a pI of 4.16. It was active against a majority of group A streptococci and displayed high levels of activity as well as binding specificity against group B and C streptococci, while it was less efficient against other streptococcal species. PlyPy showed the highest activity at neutral pH in the presence of calcium and NaCl. Surprisingly, its activity was not affected by the presence of the group A-specific carbohydrate, while the activity of PlyC was partly inhibited. Additionally, PlyPy was active in vivo and could rescue mice from systemic bacteremia. Finally, we developed a novel method to determine the peptidoglycan bond cleaved by lysins and concluded that PlyPy exhibits a rare d-alanyl-l-alanine endopeptidase activity. PlyPy thus represents the first lysin characterized from Streptococcus pyogenes and has a mechanism of action distinct from that of PlyC. PMID:24637688

  20. M-Protein Analysis of Streptococcus pyogenes Isolates Associated with Acute Rheumatic Fever in New Zealand.

    PubMed

    Williamson, Deborah A; Smeesters, Pierre R; Steer, Andrew C; Steemson, John D; Ng, Adrian C H; Proft, Thomas; Fraser, John D; Baker, Michael G; Morgan, Julie; Carter, Philip E; Moreland, Nicole J

    2015-11-01

    We applied an emm cluster typing system to group A Streptococcus strains in New Zealand, including those associated with acute rheumatic fever (ARF). We observed few so-called rheumatogenic emm types but found a high proportion of emm types previously associated with pyoderma, further suggesting a role for skin infection in ARF. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  1. Pyogenic Pericarditis and Cardiac Tamponade Due to Streptococcus anginosus in a Combat Theater

    PubMed Central

    Tripp, Michael S.; Franzos, Tracy; Wallace, Scott C.; Drinkwine, Benjamin J.; Villines, Todd C.

    2017-01-01

    Abstract Streptococcus anginosus group pericarditis is rare. A 24-year-old male soldier presented for care at a military clinic in Afghanistan with shock and cardiac tamponade requiring emergent pericardial drainage and aeromedical evacuation. We review the patient’s case, the need for serial pericardial drainage, and the available literature on this disorder. PMID:28470013

  2. Clindamycin resistant emm33 Streptococcus pyogenes emerged among invasive infections in Helsinki metropolitan area, Finland, 2012 to 2013.

    PubMed

    Pesola, A K; Sihvonen, R; Lindholm, L; Pätäri-Sampo, A

    2015-05-07

    In 2012, blood, skin and soft tissue infections caused by clindamycin resistant Streptococcus pyogenes (group A streptococcus; GAS) appeared to be increasing in the Helsinki metropolitan area. We compared monthly percentages of clindamycin resistant isolates in the area between 2012 and 2013, with those in 2010 and 2011. Resistance frequency in terms of patient age was also studied. We reviewed the medical records of bacteraemic cases in 2012 and 2013 and linked the data to emm types. To inform on the emm distribution among GAS isolated from skin and soft tissue infections during the epidemic, GAS isolates of one month (March 2013) were emm typed. For GAS blood, skin, and soft tissue isolates taken together, the proportions of clindamycin resistant isolates were significantly higher in 2012 and 2013 (23% and 17%, respectively) compared with the two previous years (3%, p<0,001). The erythromycin resistance percentages were almost equal to clindamycin (22% and 17%) in 2012 and 2013, respectively. Clindamycin resistance was most frequent in GAS isolates of 40 to 60 year-old patients (148/417; 36%). Among clindamycin resistant isolates, 12 of 14 blood isolates from 2012 to 2013, and 11 of 13 skin and soft tissue isolates from March 2013, were emm33. Emm33 GAS bacteraemia was associated with clindamycin and erythromycin resistance (odds ratio (OR): 7.0; 95% confidence interval (CI): 1.9-25.3). Infection focus was mainly the skin; either cellulitis (7/12) or necrotising fasciitis (3/12). All emm33 GAS isolates harboured the ermTR resistance gene with constitutive macrolides, lincosamides and streptogramines B (MLS(B)) phenotype. Emm33 GAS was responsible for the higher proportion of clindamycin resistance in skin, soft tissue, and blood isolates locally in 2012 and 2013.

  3. Streptococcus pyogenes infection induces septic arthritis with increased production of the receptor activator of the NF-kappaB ligand.

    PubMed

    Sakurai, Atsuo; Okahashi, Nobuo; Nakagawa, Ichiro; Kawabata, Shigetada; Amano, Atsuo; Ooshima, Takashi; Hamada, Shigeyuki

    2003-10-01

    Bacterial arthritis is a rapidly progressive and highly destructive joint disease in humans, with Staphylococcus aureus and Neisseria gonorrhoeae the major causative agents, although beta-hemolytic streptococci as well often induce the disease. We demonstrate here that intravenous inoculation of CD-1 mice with the group A streptococcus (GAS) species Streptococcus pyogenes resulted in a high incidence of septic arthritis. Signs of arthritis emerged within the first few days after injection, and bacterial examinations revealed that colonization of the inoculated GAS in the arthritic joints persisted for 21 days. Induction of persistent septic arthritis was dependent on the number of microorganisms inoculated. Immunohistochemical staining of GAS with anti-GAS antibodies revealed colonization in the joints of infected mice. Cytokine levels were quantified in the joints and sera of infected mice by using an enzyme-linked immunosorbent assay. High levels of interleukin-1beta (IL-1beta) and IL-6 were detected in the joints from 3 to 20 days after infection. We noted that an increase in the amount of receptor activator of NF-kappaB ligand (RANKL), which is a key cytokine in osteoclastogenesis, was also evident in the joints of the infected mice. RANKL was not detected in sera, indicating local production of RANKL in the infected joints. Blocking of RANKL by osteoprotegerin, a decoy receptor of RANKL, prevented bone destruction in the infected joints. These results suggest that GAS can colonize in the joints and induce bacterial arthritis. Local RANKL production in the infected joints may be involved in bone destruction.

  4. Generation and Surface Localization of Intact M Protein in Streptococcus pyogenes Are Dependent on sagA

    PubMed Central

    Biswas, Indranil; Germon, Pierre; McDade, Kathleen; Scott, June R.

    2001-01-01

    The M protein is an important surface-located virulence factor of Streptococcus pyogenes, the group A streptococcus (GAS). Expression of M protein is primarily controlled by Mga, a transcriptional activator protein. A recent report suggested that the sag locus, which includes nine genes necessary and sufficient for production of streptolysin S, another GAS virulence factor, is also needed for transcription of emm, encoding the M protein (Z. Li, D. D. Sledjeski, B. Kreikemeyer, A. Podbielski, and M. D. Boyle, J. Bacteriol. 181:6019–6027, 1999). To investigate this in more detail, we constructed an insertion-deletion mutation in sagA, the first gene in the sag locus, in the M6 strain JRS4. The resulting strain, JRS470, produced no detectable streptolysin S and showed a drastic reduction in cell surface-associated M protein, as measured by cell aggregation and Western blot analysis. However, transcription of the emm gene was unaffected by the sagA mutation. Detailed analysis with monoclonal antibodies and an antipeptide antibody showed that the M protein in the sagA mutant strain was truncated so that it lacks the C-repeat region and the C-terminal domain required for anchoring it to the cell surface. This truncated M protein was largely found, as expected, in the culture supernatant. Lack of surface-located M protein made the sagA mutant strain susceptible to phagocytosis. Thus, although sagA does not affect transcription of the M6 protein gene, it is needed for the surface localization of this important virulence factor. PMID:11598078

  5. Mode of expression and functional characterization of FCT-3 pilus region-encoded proteins in Streptococcus pyogenes serotype M49.

    PubMed

    Nakata, Masanobu; Köller, Thomas; Moritz, Karin; Ribardo, Deborah; Jonas, Ludwig; McIver, Kevin S; Sumitomo, Tomoko; Terao, Yutaka; Kawabata, Shigetada; Podbielski, Andreas; Kreikemeyer, Bernd

    2009-01-01

    The human pathogen Streptococcus pyogenes (group A streptococcus [GAS]) pilus components, suggested to play a role in pathogenesis, are encoded in the variable FCT (fibronectin- and collagen-binding T-antigen) region. We investigated the functions of sortase A (SrtA), sortase C2 (SrtC2), and the FctA protein of the most prevalent type 3 FCT region from a serotype M49 strain. Although it is considered a housekeeping sortase, SrtA's activity is involved in pilus formation in addition to its essentiality for GAS extracellular matrix protein binding, host cell adherence/internalization, survival in human blood, and biofilm formation. SrtC2 activity is crucial for pilus formation but dispensable for the other phenotypes tested in vitro. FctA is the major pilus backbone protein, simultaneously acting as the M49 T antigen, and requires SrtC2 and LepA, a signal peptidase I homologue, for monomeric surface expression and polymerization, respectively. Collagen-binding protein Cpa expression supports pilus formation at the pilus base. Immunofluorescence microscopy and fluorescence-activated cell sorting analysis revealed several unexpected expression patterns, as follows: (i) the monomeric pilus protein FctA was found exclusively at the old poles of GAS cells, (ii) FctA protein expression increased with lower temperatures, and (iii) FctA protein expression was restricted to 20 to 50% of a given GAS M49 population, suggesting regulation by a bistability mode. Notably, disruption of pilus assembly by sortase deletion rendered GAS serotype M49 significantly more aggressive in a dermonecrotic mouse infection model, indicating that sortase activity and, consequently, pilus expression allow a subpopulation of this GAS serotype to be less aggressive. Thus, pilus expression may not be a virulence attribute of GAS per se.

  6. Transcription of the Streptococcus pyogenes hyaluronic acid capsule biosynthesis operon is regulated by previously unknown upstream elements.

    PubMed

    Falaleeva, Marina; Zurek, Oliwia W; Watkins, Robert L; Reed, Robert W; Ali, Hadeel; Sumby, Paul; Voyich, Jovanka M; Korotkova, Natalia

    2014-12-01

    The important human pathogen Streptococcus pyogenes (group A Streptococcus [GAS]) produces a hyaluronic acid (HA) capsule that plays critical roles in immune evasion. Previous studies showed that the hasABC operon encoding the capsule biosynthesis enzymes is under the control of a single promoter, P1, which is negatively regulated by the two-component regulatory system CovR/S. In this work, we characterize the sequence upstream of P1 and identify a novel regulatory region controlling transcription of the capsule biosynthesis operon in the M1 serotype strain MGAS2221. This region consists of a promoter, P2, which initiates transcription of a novel small RNA, HasS, an intrinsic transcriptional terminator that inefficiently terminates HasS, permitting read-through transcription of hasABC, and a putative promoter which lies upstream of P2. Electrophoretic mobility shift assays, quantitative reverse transcription-PCR, and transcriptional reporter data identified CovR as a negative regulator of P2. We found that the P1 and P2 promoters are completely repressed by CovR, and capsule expression is regulated by the putative promoter upstream of P2. Deletion of hasS or of the terminator eliminates CovR-binding sequences, relieving repression and increasing read-through, hasA transcription, and capsule production. Sequence analysis of 44 GAS genomes revealed a high level of polymorphism in the HasS sequence region. Most of the HasS variations were located in the terminator sequences, suggesting that this region is under strong selective pressure. We discovered that the terminator deletion mutant is highly resistant to neutrophil-mediated killing and is significantly more virulent in a mouse model of GAS invasive disease than the wild-type strain. Together, these results are consistent with the naturally occurring mutations in this region modulating GAS virulence.

  7. Preliminary validation of a novel high-resolution melt-based typing method based on the multilocus sequence typing scheme of Streptococcus pyogenes.

    PubMed

    Richardson, L J; Tong, S Y C; Towers, R J; Huygens, F; McGregor, K; Fagan, P K; Currie, B J; Carapetis, J R; Giffard, P M

    2011-09-01

    The major limitation of current typing methods for Streptococcus pyogenes, such as emm sequence typing and T typing, is that these are based on regions subject to considerable selective pressure. Multilocus sequence typing (MLST) is a better indicator of the genetic backbone of a strain but is not widely used due to high costs. The objective of this study was to develop a robust and cost-effective alternative to S. pyogenes MLST. A 10-member single nucleotide polymorphism (SNP) set that provides a Simpson's Index of Diversity (D) of 0.99 with respect to the S. pyogenes MLST database was derived. A typing format involving high-resolution melting (HRM) analysis of small fragments nucleated by each of the resolution-optimized SNPs was developed. The fragments were 59-119 bp in size and, based on differences in G+C content, were predicted to generate three to six resolvable HRM curves. The combination of curves across each of the 10 fragments can be used to generate a melt type (MelT) for each sequence type (ST). The 525 STs currently in the S. pyogenes MLST database are predicted to resolve into 298 distinct MelTs and the method is calculated to provide a D of 0.996 against the MLST database. The MelTs are concordant with the S. pyogenes population structure. To validate the method we examined clinical isolates of S. pyogenes of 70 STs. Curves were generated as predicted by G+C content discriminating the 70 STs into 65 distinct MelTs.

  8. Chemokine-cleaving Streptococcus pyogenes protease SpyCEP is necessary and sufficient for bacterial dissemination within soft tissues and the respiratory tract.

    PubMed

    Kurupati, Prathiba; Turner, Claire E; Tziona, Ioanna; Lawrenson, Richard A; Alam, Faraz M; Nohadani, Mahrokh; Stamp, Gordon W; Zinkernagel, Annelies S; Nizet, Victor; Edwards, Robert J; Sriskandan, Shiranee

    2010-06-01

    SpyCEP is a Streptococcus pyogenes protease that cleaves CXCL8/IL-8 and its activity is associated with human invasive disease severity. We investigated the role of SpyCEP in S. pyogenes necrotizing fasciitis and respiratory tract infection in mice using isogenic strains differing only in SpyCEP expression. SpyCEP cleaved human CXCL1, 2, 6 and 8 plus murine CXCL1 and 2 at a structurally conserved site. Mice were infected in thigh muscle with a strain of S. pyogenes that expresses a high level of SpyCEP, or with an isogenic non-SpyCEP expressing strain. SpyCEP expression by S. pyogenes hindered bacterial clearance from muscle, and enhanced bacterial spread, associated with cleavage of murine chemoattractant CXCL1. Mice were then infected with Lactococcus lactis strains that differed only in SpyCEP expression. In contrast to the parent L. lactis strain (lacks SpyCEP), which was avirulent when administered intramuscularly, infection with a strain that expressed SpyCEP heterologously led to dramatic systemic illness within 24 h, failure to clear bacteria from muscle and marked dissemination to other organs. In the upper airways, SpyCEP expression was required for survival of L. lactis but not S. pyogenes. However, dissemination of S. pyogenes to the lung was SpyCEP-dependent and was associated with evidence of chemokine cleavage. Taken together, the studies provide clear evidence that SpyCEP is necessary and sufficient for systemic bacterial dissemination from a soft tissue focus in this model and also underlies dissemination in the respiratory tract.

  9. Chemokine-cleaving Streptococcus pyogenes protease SpyCEP is necessary and sufficient for bacterial dissemination within soft tissues and the respiratory tract

    PubMed Central

    Kurupati, Prathiba; Turner, Claire E; Tziona, Ioanna; Lawrenson, Richard A; Alam, Faraz M; Nohadani, Mahrokh; Stamp, Gordon W; Zinkernagel, Annelies S; Nizet, Victor; Edwards, Robert J; Sriskandan, Shiranee

    2010-01-01

    SpyCEP is a Streptococcus pyogenes protease that cleaves CXCL8/IL-8 and its activity is associated with human invasive disease severity. We investigated the role of SpyCEP in S. pyogenes necrotizing fasciitis and respiratory tract infection in mice using isogenic strains differing only in SpyCEP expression. SpyCEP cleaved human CXCL1, 2, 6 and 8 plus murine CXCL1 and 2 at a structurally conserved site. Mice were infected in thigh muscle with a strain of S. pyogenes that expresses a high level of SpyCEP, or with an isogenic non-SpyCEP expressing strain. SpyCEP expression by S. pyogenes hindered bacterial clearance from muscle, and enhanced bacterial spread, associated with cleavage of murine chemoattractant CXCL1. Mice were then infected with Lactococcus lactis strains that differed only in SpyCEP expression. In contrast to the parent L. lactis strain (lacks SpyCEP), which was avirulent when administered intramuscularly, infection with a strain that expressed SpyCEP heterologously led to dramatic systemic illness within 24 h, failure to clear bacteria from muscle and marked dissemination to other organs. In the upper airways, SpyCEP expression was required for survival of L. lactis but not S. pyogenes. However, dissemination of S. pyogenes to the lung was SpyCEP-dependent and was associated with evidence of chemokine cleavage. Taken together, the studies provide clear evidence that SpyCEP is necessary and sufficient for systemic bacterial dissemination from a soft tissue focus in this model and also underlies dissemination in the respiratory tract. PMID:20158613

  10. Streptococcus pyogenes Endopeptidase O Contributes to Evasion from Complement-mediated Bacteriolysis via Binding to Human Complement Factor C1q.

    PubMed

    Honda-Ogawa, Mariko; Sumitomo, Tomoko; Mori, Yasushi; Hamd, Dalia Talat; Ogawa, Taiji; Yamaguchi, Masaya; Nakata, Masanobu; Kawabata, Shigetada

    2017-03-10

    Streptococcus pyogenes secretes various virulence factors for evasion from complement-mediated bacteriolysis. However, full understanding of the molecules possessed by this organism that interact with complement C1q, an initiator of the classical complement pathway, remains elusive. In this study, we identified an endopeptidase of S. pyogenes, PepO, as an interacting molecule, and investigated its effects on complement immunity and pathogenesis. Enzyme-linked immunosorbent assay and surface plasmon resonance analysis findings revealed that S. pyogenes recombinant PepO bound to human C1q in a concentration-dependent manner under physiological conditions. Sites of inflammation are known to have decreased pH levels, thus the effects of PepO on bacterial evasion from complement immunity was analyzed in a low pH condition. Notably, under low pH conditions, PepO exhibited a higher affinity for C1q as compared with IgG, and PepO inhibited the binding of IgG to C1q. In addition, pepO deletion rendered S. pyogenes more susceptible to the bacteriocidal activity of human serum. Also, observations of the morphological features of the pepO mutant strain (ΔpepO) showed damaged irregular surfaces as compared with the wild-type strain (WT). WT-infected tissues exhibited greater severity and lower complement activity as compared with those infected by ΔpepO in a mouse skin infection model. Furthermore, WT infection resulted in a larger accumulation of C1q than that with ΔpepO. Our results suggest that interaction of S. pyogenes PepO with C1q interferes with the complement pathway, which enables S. pyogenes to evade complement-mediated bacteriolysis under acidic conditions, such as seen in inflammatory sites. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. Antimicrobial Susceptibilities of 1,684 Streptococcus pneumoniae and 2,039 Streptococcus pyogenes Isolates and Their Ecological Relationships: Results of a 1-Year (1998–1999) Multicenter Surveillance Study in Spain

    PubMed Central

    Pérez-Trallero, E.; Fernández-Mazarrasa, C.; García-Rey, C.; Bouza, E.; Aguilar, L.; García-de-Lomas, J.; Baquero, F.

    2001-01-01

    A nationwide multicenter susceptibility surveillance study which included 1,684 Streptococcus pneumoniae and 2,039 S. pyogenes isolates was carried out over 1 year in order to assess the current resistance patterns for the two most important gram-positive microorganisms responsible for community-acquired infections in Spain. Susceptibility testing was done by a broth microdilution method according to National Committee for Clinical Laboratory Standards M100-S10 interpretative criteria. For S. pneumoniae, the prevalences of highly resistant strains were 5% for amoxicillin and amoxicillin-clavulanic acid; 7% for cefotaxime; 22% for penicillin; 31% for cefuroxime; 35% for erythromycin, clarithromycin, and azithromycin; and 42% for cefaclor. For S. pyogenes, the prevalence of erythromycin resistance was 20%. Efflux was encountered in 90% of S. pyogenes and 5% of S. pneumoniae isolates that exhibited erythromycin resistance. Erythromycin resistance was associated with clarithromycin and azithromycin in both species, regardless of phenotype. Despite the different nature of the mechanisms of resistance, a positive correlation (r = 0.612) between the two species in the prevalence of erythromycin resistance was found in site-by-site comparisons, suggesting some kind of link with antibiotic consumption. Regarding ciprofloxacin, the MIC was ≥4 μg/ml for 7% of S. pneumoniae and 3.5% of S. pyogenes isolates. Ciprofloxacin resistance (MIC, ≥4 μg/ml) was significantly (P < 0.05) associated with macrolide resistance in both S. pyogenes and S. pneumoniae and with penicillin nonsusceptibility in S. pneumoniae. PMID:11709305

  12. Inactivation of Streptococcus pyogenes extracellular cysteine protease significantly decreases mouse lethality of serotype M3 and M49 strains.

    PubMed Central

    Lukomski, S; Sreevatsan, S; Amberg, C; Reichardt, W; Woischnik, M; Podbielski, A; Musser, J M

    1997-01-01

    Cysteine proteases have been implicated as important virulence factors in a wide range of prokaryotic and eukaryotic pathogens, but little direct evidence has been presented to support this notion. Virtually all strains of the human bacterial pathogen Streptococcus pyogenes express a highly conserved extracellular cysteine protease known as streptococcal pyrogenic exotoxin B (SpeB). Two sets of isogenic strains deficient in SpeB cysteine protease activity were constructed by integrational mutagenesis using nonreplicating recombinant plasmids containing a truncated segment of the speB gene. Immunoblot analyses and enzyme assays confirmed that the mutant derivatives were deficient in expression of enzymatically active SpeB cysteine protease. To test the hypothesis that the cysteine protease participates in host mortality, we assessed the ability of serotype M3 and M49 wild-type strains and isogenic protease-negative mutants to cause death in outbred mice after intraperitoneal inoculation. Compared to wild-type parental organisms, the serotype M3 speB mutant lost virtually all ability to cause mouse death (P < 0.00001), and similarly, the virulence of the M49 mutant was detrimentally altered (P < 0.005). The data unambiguously demonstrate that the streptococcal enzyme is a virulence factor, and thereby provide additional evidence that microbial cysteine proteases are critical in host-pathogen interactions. PMID:9169486

  13. Characterisation of the main clones of Streptococcus pyogenes carrying the ermA (subclass TR) gene in Spain.

    PubMed

    Montes, Milagrosa; Orden, Beatriz; Tamayo, Esther; Alos, Juan-Ignacio; Pérez-Trallero, Emilio

    2006-11-01

    Seventy-four Streptococcus pyogenes isolates showing the macrolide-lincosamide-streptogramin B (MLS(B)) resistance phenotype carrying the ermA gene (72 of which showed the inducible resistance phenotype) were obtained between 1999 and 2004. Seven different sequence types (STs) and emm types were detected: emm22/ST46 (n=33); emm77/ST63 (n=22); emm73/ST331 (n=10); emm94/ST89 (n=6); and one isolate each of emm28/ST52, emm11/ST403 and emm4/ST38. All ST46 isolates were susceptible to tetracycline and almost all reacted against the T12 type (all agglutinated into the T-pattern 3/12/13/B3264). Resistance to tetracycline was observed in all ST63 (tetO+) and ST89 (tetM+) isolates. Most of the ST63 isolates reacted against the T28 type (all agglutinated into the T-pattern 9/13/28). The 74 isolates were grouped into eight pulsed-field gel electrophoresis pulsotypes (one cluster for each emm/ST type, except for emm77/ST63).

  14. One-pot enzymatic glycan remodeling of a therapeutic monoclonal antibody by endoglycosidase S (Endo-S) from Streptococcus pyogenes.

    PubMed

    Tong, Xin; Li, Tiezheng; Orwenyo, Jared; Toonstra, Christian; Wang, Lai-Xi

    2017-07-29

    A facile, one-pot enzymatic glycan remodeling of antibody rituximab to produce homogeneous high-mannose and hybrid type antibody glycoforms is described. This method was based on the unique substrate specificity of the endoglycosidase S (Endo-S) from Streptococcus pyogenes. While Endo-S efficiently hydrolyzes the bi-antennary complex type IgG Fc N-glycans, we found that Endo-S did not hydrolyze the "ground state" high-mannose or hybrid glycoforms, and only slowly hydrolyzed the highly activated high-mannose or hybrid N-glycan oxazolines. Moreover, we found that wild-type Endo-S could efficiently use high-mannose or hybrid glycan oxazolines for transglycosylation without product hydrolysis. The combination of the remarkable difference in substrate specificity of Endo-S allows the deglycosylation of heterogeneous rituximab and the transglycosylation with glycan oxazoline to take place in one-pot without the need of isolating the deglycosylated intermediate or changing the enzyme to afford the high-mannose type, hybrid type, and some selectively modified truncated form of antibody glycoforms. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. The AgI/II family adhesin AspA is required for respiratory infection by Streptococcus pyogenes.

    PubMed

    Franklin, Linda; Nobbs, Angela H; Bricio-Moreno, Laura; Wright, Christopher J; Maddocks, Sarah E; Sahota, Jaspreet Singh; Ralph, Joe; O'Connor, Matthew; Jenkinson, Howard F; Kadioglu, Aras

    2013-01-01

    Streptococcus pyogenes (GAS) is a human pathogen that causes pharyngitis and invasive diseases such as toxic shock syndrome and sepsis. The upper respiratory tract is the primary reservoir from which GAS can infect new hosts and cause disease. The factors involved in colonisation are incompletely known however. Previous evidence in oral streptococci has shown that the AgI/II family proteins are involved. We hypothesized that the AspA member of this family might be involved in GAS colonization. We describe a novel mouse model of GAS colonization of the nasopharynx and lower respiratory tract to elucidate these interactions. We used two clinical M serotypes expressing AspA, and their aspA gene deletant isogenic mutants in experiments using adherence assays to respiratory epithelium, macrophage phagocytosis and neutrophil killing assays and in vivo models of respiratory tract colonisation and infection. We demonstrated the requirement for AspA in colonization of the respiratory tract. AspA mutants were cleared from the respiratory tract and were deficient in adherence to epithelial cells, and susceptible to phagocytosis. Expression of AspA in the surrogate host Lactococcus lactis protected bacteria from phagocytosis. Our results suggest that AspA has an essential role in respiratory infection, and may function as a novel anti-phagocytic factor.

  16. Identification of residues responsible for the defective virulence gene regulator Mga produced by a natural mutant of Streptococcus pyogenes.

    PubMed

    Vahling, Cheryl M; McIver, Kevin S

    2005-09-01

    Mga is a transcriptional regulator in the pathogen Streptococcus pyogenes that positively activates several important virulence genes involved in colonization and immune evasion in the human host. A naturally occurring mutant of Mga that is defective in its ability to activate transcription has been identified in the serotype M50 strain B514-Sm. Sequence alignment of the defective M50 Mga with the fully functional Mga from serotypes M4 and M49 revealed only three amino acid changes that might result in a defective protein. Electrophoretic mobility shift assays using purified M50 and M4 maltose binding protein-Mga found that both exhibited DNA-binding activity towards regulated promoters. Thus, the significance of each residue for the functionality of M50 Mga was explored through introduction of "gain-of-function" mutations based on M4 Mga. Transcriptional studies of the mutant alleles under both constitutive (PrpsL) and autoactivated (Pmga4) promoters illustrated that an arginine-to-methionine change at position 461 of M50 Mga protein fully restored activation of downstream genes. Western blot analyses of steady-state Mga levels suggest that the M461 residue may play a role in overall conformation and protein stability of Mga. However, despite the conservation of the M461 protein among all other Mga proteins, it does not appear to be necessary for activity in a divergent M6 Mga. These studies highlight the potential differences that exist between divergent Mga proteins in this important human pathogen.

  17. Streptococcus pyogenes Sortase Mutants Are Highly Susceptible to Killing by Host Factors Due to Aberrant Envelope Physiology

    PubMed Central

    Raz, Assaf; Tanasescu, Ana-Maria; Zhao, Anna M.; Serrano, Anna; Alston, Tricia; Sol, Asaf; Bachrach, Gilad; Fischetti, Vincent A.

    2015-01-01

    Cell wall anchored virulence factors are critical for infection and colonization of the host by Gram-positive bacteria. Such proteins have an N-terminal leader sequence and a C-terminal sorting signal, composed of an LPXTG motif, a hydrophobic stretch, and a few positively charged amino acids. The sorting signal halts translocation across the membrane, allowing sortase to cleave the LPXTG motif, leading to surface anchoring. Deletion of sortase prevents the anchoring of virulence factors to the wall; the effects on bacterial physiology however, have not been thoroughly characterized. Here we show that deletion of Streptococcus pyogenes sortase A leads to accumulation of sorting intermediates, particularly at the septum, altering cellular morphology and physiology, and compromising membrane integrity. Such cells are highly sensitive to cathelicidin, and are rapidly killed in blood and plasma. These phenomena are not a loss-of-function effect caused by the absence of anchored surface proteins, but specifically result from the accumulation of sorting intermediates. Reduction in the level of sorting intermediates leads to a return of the sortase mutant to normal morphology, while expression of M protein with an altered LPXTG motif in wild type cells leads to toxicity in the host environment, similar to that observed in the sortase mutant. These unanticipated effects suggest that inhibition of sortase by small-molecule inhibitors could similarly lead to the rapid elimination of pathogens from an infected host, making such inhibitors much better anti-bacterial agents than previously believed. PMID:26484774

  18. [Resistance to macrolides in the species Streptococcus pyogenes in the Czech Republic in 1996-2003].

    PubMed

    Urbásková, P; Jakubů, V

    2004-11-01

    The study of the prevalence of erythromycin resistance in 22 169 S. pyogenes strains in the Czech Republic in 1996-2003 on the background of rough data on the nationwide consumption of macrolide antibiotics confirmed that the exponential growth of resistance observed in 1998-2001 copied with a delay the rise in macrolide antibiotic consumption recorded in 1992-1995. The highest frequency of erythromycin resistance was found in 2001 (16.5%) with a subsequent decrease to 14.5% in 2002 and to 9.1% in 2003. The drop in resistance followed the stagnation in macrolide consumption and its decrease by 17% in 2002. Upward and downward trends in macrolide resistance in different regions and age groups copied the nationwide trends with some quantitative differences that could not be analyzed in view of the lack of detailed data on antibiotic consumption. A 99.5% concordance was found between the results of the phenotypic method and those of detection of genes coding for constitutive, inducible and efflux resistance to macrolide-lincosamide-streptograminB (MLSB) antibiotics. In 2001 when the highest erythromycin resistance was recorded in the Czech Republic, most of the tested strains (91.2%) showed resistance to all MLSB antibiotics, with macrolide efflux (susceptibility to lincosamides and 16-membered macrolides was conserved) being implicated in resistance of 8.8% of the strains only. In 2003, the number of erythromycin resistant strains decreased and the resistance mechanism was ascribed to macrolide efflux in 26.8% of them. Almost all of the strains with constitutive or induced MLSB resistance are also resistant to either tetracycline or bacitracin or both. In the light of S. pyogenes resistance to bacitracin, the bacitracin disk is not usable in preliminary identification any more.

  19. Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of SpyCEP, a candidate antigen for a vaccine against Streptococcus pyogenes.

    PubMed

    Abate, Francesca; Malito, Enrico; Falugi, Fabiana; Margarit Y Ros, Immaculada; Bottomley, Matthew James

    2013-10-01

    Streptococcus pyogenes (Group A streptococcus; GAS) is an important human pathogen against which an effective vaccine does not yet exist. The S. pyogenes protein SpyCEP (S. pyogenes cell-envelope proteinase) is a surface-exposed subtilisin-like serine protease of 1647 amino acids. In addition to its auto-protease activity, SpyCEP is capable of cleaving interleukin 8 and related chemokines, contributing to GAS immune-evasion strategies. SpyCEP is immunogenic and confers protection in animal models of GAS infections. In order to structurally characterize this promising vaccine candidate, several SpyCEP protein-expression constructs were designed, cloned, produced in Escherichia coli, purified by affinity chromatography and subjected to crystallization trials. Crystals of a selenomethionyl form of a near-full-length SpyCEP ectodomain were obtained. The crystals diffracted X-rays to 3.3 Å resolution and belonged to space group C2, with unit-cell parameters a=139.2, b=120.4, c=104.3 Å, β=111°.

  20. Potential antibiotic and anti-infective effects of rhodomyrtone from Rhodomyrtus tomentosa (Aiton) Hassk. on Streptococcus pyogenes as revealed by proteomics.

    PubMed

    Limsuwan, Surasak; Hesseling-Meinders, Anne; Voravuthikunchai, Supayang Piyawan; van Dijl, Jan Maarten; Kayser, Oliver

    2011-08-15

    Rhodomyrtone from Rhodomyrtus tomentosa (Aiton) Hassk. leaf extract has a strong antibacterial activity against the bacterial pathogen Streptococcus pyogenes. Our previous studies indicated that the bactericidal activity of rhodomyrtone might involve intracellular targets. In the present studies we followed a proteomics approach to investigate the mode of action of rhodomyrtone on S. pyogenes. For this purpose, S. pyogenes was cultivated in the presence of 0.39 μg/ml rhodomyrtone, which corresponds to 50% of the minimal inhibitory concentration. The results show that the amounts of various enzymes associated with important metabolic pathways were strongly affected, which is consistent with the growth-inhibiting effect of rhodomyrtone. Additionally, cells of S. pyogenes grown in the presence of rhodomyrtone produced reduced amounts of known virulence factors, such as the glyceraldehyde-3-phosphate dehydrogenase, the CAMP factor, and the streptococcal pyrogenic exotoxin C. Taken together, these findings indicate that rhodomyrtone has both antimicrobial and anti-infective activities, which make it an interesting candidate drug.

  1. Inactivation of the CovR/S Virulence Regulator Impairs Infection in an Improved Murine Model of Streptococcus pyogenes Naso-Pharyngeal Infection

    PubMed Central

    Alam, Faraz M.; Turner, Claire E.; Smith, Ken; Wiles, Siouxsie; Sriskandan, Shiranee

    2013-01-01

    Streptococcus pyogenes is a leading cause of pharyngeal infection, with an estimated 616 million cases per year. The human nasopharynx represents the major reservoir for all S. pyogenes infection, including severe invasive disease. To investigate bacterial and host factors that influence S. pyogenes infection, we have devised an improved murine model of nasopharyngeal colonization, with an optimized dosing volume to avoid fulminant infections and a sensitive host strain. In addition we have utilized a refined technique for longitudinal monitoring of bacterial burden that is non-invasive thereby reducing the numbers of animals required. The model was used to demonstrate that the two component regulatory system, CovR/S, is required for optimum infection and transmission from the nasopharynx. There is a fitness cost conferred by covR/S mutation that is specific to the nasopharynx. This may explain why S. pyogenes with altered covR/S have not become prevalent in community infections despite possessing a selective advantage in invasive infection. PMID:23637876

  2. Comparative genomics of Streptococcus pyogenes M1 isolates differing in virulence and propensity to cause systemic infection in mice.

    PubMed

    Fiebig, Anne; Loof, Torsten G; Babbar, Anshu; Itzek, Andreas; Koehorst, Jasper J; Schaap, Peter J; Nitsche-Schmitz, D Patric

    2015-09-01

    Streptococcus pyogenes serotype M1 is a frequent cause of severe infections in humans. Some M1 isolates are pathogenic in mice and used in studies on infection pathogenesis. We observed marked differences in murine infections caused by M1 strain SF370, 5448, 5448AP or AP1 which prompted us to sequence the whole genome of isolates 5448 and AP1 for comparative analysis. Strain 5448 is known to acquire inactivating mutations in the CovRS two-component system during mouse infection, producing hypervirulent progeny such as 5448AP. Isolates AP1 and 5448AP, more than 5448, caused disseminating infections that became systemic and lethal. SF370 was not pathogenic. Phages caused gross genetic differences and increased the gene content of AP1 by 8% as compared to 5448 and SF370. Each of six examined M1 genomes contained two CRISPR-Cas systems. Phage insertion destroyed a type II CRISPR-Cas system in AP1 and other strains of serotypes M1, M3, M6 and M24, but not in M1 strains 5448, SF370, MGAS5005, A20 or M1 476. A resulting impaired defence against invading genetic elements could have led to the wealth of phages in AP1. AP1 lacks genetic features of the MGAS5005-like clonal complex including the streptodornase that drives selection for hypervirulent clones with inactivated CovRS system. Still, inactivating mutations in covS were a common genetic feature of AP1 and the MGAS5005-like isolate 5448AP. Abolished expression of the cysteine proteinase SpeB, due to CovRS inactivation could be a common cause for hypervirulence of the two isolates. Moreover, an additional protein H-coding gene and a mutation in the regulator gene rofA distinguished AP1 form other M1 isolates. In conclusion, hypervirulence of S. pyogenes M1 in mice is not limited to the MGAS5005-like genotype. Copyright © 2015 Elsevier GmbH. All rights reserved.

  3. Genomic Characterization of a Pattern D Streptococcus pyogenes emm53 Isolate Reveals a Genetic Rationale for Invasive Skin Tropicity.

    PubMed

    Bao, Yun-Juan; Liang, Zhong; Mayfield, Jeffrey A; Donahue, Deborah L; Carothers, Katelyn E; Lee, Shaun W; Ploplis, Victoria A; Castellino, Francis J

    2016-06-15

    The genome of an invasive skin-tropic strain (AP53) of serotype M53 group A Streptococcus pyogenes (GAS) is composed of a circular chromosome of 1,860,554 bp and carries genetic markers for infection at skin locales, viz, emm gene family pattern D and FCT type 3. Through genome-scale comparisons of AP53 with other GAS genomes, we identified 596 candidate single-nucleotide polymorphisms (SNPs) that reveal a potential genetic basis for skin tropism. The genome of AP53 differed by ∼30 point mutations from a noninvasive pattern D serotype M53 strain (Alab49), 4 of which are located in virulence genes. One pseudogene, yielding an inactive sensor kinase (CovS(-)) of the two-component transcriptional regulator CovRS, a major determinant for invasiveness, severely attenuated the expression of the secreted cysteine protease SpeB and enhanced the expression of the hyaluronic acid capsule compared to the isogenic noninvasive AP53/CovS(+) strain. The collagen-binding protein transcript sclB differed in the number of 5'-pentanucleotide repeats in the signal peptides of AP53 and Alab49 (9 versus 15), translating into different lengths of their signal peptides, which nonetheless maintained a full-length translatable coding frame. Furthermore, GAS strain AP53 acquired two phages that are absent in Alab49. One such phage (ΦAP53.2) contains the known virulence factor superantigen exotoxin gene tandem speK-slaA Overall, we conclude that this bacterium has evolved in multiple ways, including mutational variations of regulatory genes, short-tandem-repeat polymorphisms, large-scale genomic alterations, and acquisition of phages, all of which may be involved in shaping the adaptation of GAS in specific infectious environments and contribute to its enhanced virulence. Infectious strains of S. pyogenes (GAS) are classified by their serotypes, relating to the surface M protein, the emm-like subfamily pattern, and their tropicity toward the nasopharynx and/or skin. It is generally agreed

  4. Genomic Characterization of a Pattern D Streptococcus pyogenes emm53 Isolate Reveals a Genetic Rationale for Invasive Skin Tropicity

    PubMed Central

    Bao, Yun-Juan; Liang, Zhong; Mayfield, Jeffrey A.; Donahue, Deborah L.; Carothers, Katelyn E.; Lee, Shaun W.; Ploplis, Victoria A.

    2016-01-01

    ABSTRACT The genome of an invasive skin-tropic strain (AP53) of serotype M53 group A Streptococcus pyogenes (GAS) is composed of a circular chromosome of 1,860,554 bp and carries genetic markers for infection at skin locales, viz., emm gene family pattern D and FCT type 3. Through genome-scale comparisons of AP53 with other GAS genomes, we identified 596 candidate single-nucleotide polymorphisms (SNPs) that reveal a potential genetic basis for skin tropism. The genome of AP53 differed by ∼30 point mutations from a noninvasive pattern D serotype M53 strain (Alab49), 4 of which are located in virulence genes. One pseudogene, yielding an inactive sensor kinase (CovS−) of the two-component transcriptional regulator CovRS, a major determinant for invasiveness, severely attenuated the expression of the secreted cysteine protease SpeB and enhanced the expression of the hyaluronic acid capsule compared to the isogenic noninvasive AP53/CovS+ strain. The collagen-binding protein transcript sclB differed in the number of 5′-pentanucleotide repeats in the signal peptides of AP53 and Alab49 (9 versus 15), translating into different lengths of their signal peptides, which nonetheless maintained a full-length translatable coding frame. Furthermore, GAS strain AP53 acquired two phages that are absent in Alab49. One such phage (ΦAP53.2) contains the known virulence factor superantigen exotoxin gene tandem speK-slaA. Overall, we conclude that this bacterium has evolved in multiple ways, including mutational variations of regulatory genes, short-tandem-repeat polymorphisms, large-scale genomic alterations, and acquisition of phages, all of which may be involved in shaping the adaptation of GAS in specific infectious environments and contribute to its enhanced virulence. IMPORTANCE Infectious strains of S. pyogenes (GAS) are classified by their serotypes, relating to the surface M protein, the emm-like subfamily pattern, and their tropicity toward the nasopharynx and/or skin

  5. Toward new therapeutics for skin and soft tissue infections: propargyl-linked antifolates are potent inhibitors of MRSA and Streptococcus pyogenes.

    PubMed

    Viswanathan, Kishore; Frey, Kathleen M; Scocchera, Eric W; Martin, Brooke D; Swain Iii, P Whitney; Alverson, Jeremy B; Priestley, Nigel D; Anderson, Amy C; Wright, Dennis L

    2012-01-01

    Hospital- and community-acquired, complicated skin and soft tissue infections, often attributed to Staphylococcus aureus and Streptococcus pyogenes, present a significant health burden that is associated with increased health care costs and mortality. As these two species are difficult to discern on diagnosis and are associated with differential profiles of drug resistance, the development of an efficacious antibacterial agent that targets both organisms is a high priority. Herein we describe a structure-based drug development effort that has produced highly potent inhibitors of dihydrofolate reductase from both species. Optimized propargyl-linked antifolates containing a key pyridyl substituent display antibacterial activity against both methicillin-resistant S. aureus and S. pyogenes at MIC values below 0.1 µg/mL and minimal cytotoxicity against mammalian cells. Further evaluation against a panel of clinical isolates shows good efficacy against a range of important phenotypes such as hospital- and community-acquired strains as well as strains resistant to vancomycin.

  6. Engineering multiple biological functional motifs into a blank collagen-like protein template from Streptococcus pyogenes.

    PubMed

    Peng, Yong Y; Stoichevska, Violet; Schacht, Kristin; Werkmeister, Jerome A; Ramshaw, John A M

    2014-07-01

    Bacterially derived triple-helical, collagen-like proteins are attractive as potential biomedical materials. The collagen-like domain of the Scl2 protein from S. pyogenes lacks any specific binding sites for mammalian cells yet possesses the inherent structural integrity of the collagen triple-helix of animal collagens. It can, therefore, be considered as a structurally-stable "blank slate" into which various defined, biological sequences, derived from animal collagens, can be added by substitutions or insertions, to enable production of novel designed materials to fit specific functional requirements. In the present study, we have used site directed mutagenesis to substitute two functional sequences, one for heparin binding and the other for integrin binding, into different locations in the triple-helical structure. This provided three new constructs, two containing the single substitutions and one containing both substitutions. The stability of these constructs was marginally reduced when compared to the unmodified sequence. When compared to the unmodified bacterial collagen, both the modified collagens that contain the heparin binding site showed marked binding of fluorescently labeled heparin. Similarly, the modified collagens from both constructs containing the integrin binding site showed significant adhesion of L929 cells that are known to possess the appropriate integrin receptor. C2C12 cells that lack any appropriate integrins did not bind. These data show that bacterial collagen-like sequences can be modified to act like natural extracellular matrix collagens by inserting one or more unique biological domains with defined function.

  7. Expression profile of BAFF in peripheral blood from patients of IgA nephropathy: Correlation with clinical features and Streptococcus pyogenes infection.

    PubMed

    Zheng, Nuoyan; Fan, Jinjin; Wang, Bing; Wang, Dongxian; Feng, Pinning; Yang, Qiongqiong; Yu, Xueqing

    2017-04-01

    B cells are critically important for the pathogenesis of IgA nephropathy (IgAN). The present study aimed to investigate the abundance of B cell activating factor (BAFF), which belongs to the tumor necrosis factor superfamily, in the peripheral blood of patients with IgAN. The different forms of BAFF in peripheral blood and its association with clinical features and immunological factors were analyzed. mRNA levels of BAFF and other associated genes in the peripheral blood mononuclear cells (PBMCs) of patients with IgAN and controls were analyzed by quantitative polymerase chain reaction. Cellular BAFF proteins in PBMCs and plasma soluble BAFF proteins were measured by western blot analysis and ELISA, respectively. PBMCs from patients were stimulated with Streptococcus pyogenes (S. pyogenes) ex vivo for the BAFF secretion assay. The data demonstrated that, although mRNA levels of BAFF in PBMC were not significantly increased in patients with IgAN, they were positively associated with those of a proliferation inducing ligand (APRIL), Toll‑like receptor (TLR)2, TLR4 and TLR7. The cellular BAFF protein in PBMCs was not upregulated. Plasma BAFF protein levels in patients with IgAN (n=76) were significantly decreased compared with controls. However, plasma BAFF levels were positively associated with serum creatinine, proteinuria, uric acid and group A Streptococcus infection index in patients with IgAN. In patients with IgAN, plasma BAFF concentrations were markedly higher in those with more severe renal tubular atrophy/interstitial fibrosis and global glomerulosclerosis. Furthermore, BAFF production in PBMCs of patients with IgAN was increased following S. pyogenes stimulation ex vivo. In conclusion, plasma BAFF levels in patients with IgAN were associated with renal function and disease activity. S. pyogenes infection was closely associated with BAFF production in patients with IgAN.

  8. Streptococcus pyogenes antigen I/II-family polypeptide AspA shows differential ligand-binding properties and mediates biofilm formation.

    PubMed

    Maddocks, Sarah E; Wright, Christopher J; Nobbs, Angela H; Brittan, Jane L; Franklin, Linda; Strömberg, Nicklas; Kadioglu, Aras; Jepson, Mark A; Jenkinson, Howard F

    2011-08-01

    The streptococcal antigen I/II (AgI/II)-family polypeptides are cell wall-anchored adhesins expressed by most indigenous oral streptococci. Proteins sharing 30-40% overall amino acid sequence similarities with AgI/II-family proteins are also expressed by Streptococcus pyogenes. The S. pyogenes M28_Spy1325 polypeptide (designated AspA) displays an AgI/II primary structure, with alanine-rich (A) and proline-rich (P) repeats flanking a V region that is projected distal from the cell. In this study it is shown that AspA from serotype M28 S. pyogenes, when expressed on surrogate host Lactococcus lactis, confers binding to immobilized salivary agglutinin gp-340. This binding was blocked by antibodies to the AspA-VP region. In contrast, the N-terminal region of AspA was deficient in binding fluid-phase gp-340, and L. lactis cells expressing AspA were not agglutinated by gp-340. Deletion of the aspA gene from two different M28 strains of S. pyogenes abrogated their abilities to form biofilms on saliva-coated surfaces. In each mutant strain, biofilm formation was restored by trans complementation of the aspA deletion. In addition, expression of AspA protein on the surface of L. lactis conferred biofilm-forming ability. Taken collectively, the results provide evidence that AspA is a biofilm-associated adhesin that may function in host colonization by S. pyogenes. © 2011 Blackwell Publishing Ltd.

  9. Arginine deprivation by arginine deiminase of Streptococcus pyogenes controls primary glioblastoma growth in vitro and in vivo.

    PubMed

    Fiedler, Tomas; Strauss, Madlen; Hering, Silvio; Redanz, Ulrike; William, Doreen; Rosche, Yvonne; Classen, Carl Friedrich; Kreikemeyer, Bernd; Linnebacher, Michael; Maletzki, Claudia

    2015-01-01

    Arginine auxotrophy constitutes a weak point of several tumors, among them glioblastoma multiforme (GBM). Hence, those tumors are supposed to be sensitive for arginine-depleting substances, such as arginine deiminase (ADI). Here we elucidated the sensitivity of patient-individual GBM cell lines toward Streptococcus pyogenes-derived ADI. To improve therapy, ADI was combined with currently established and pre-clinical cytostatic drugs. Additionally, effectiveness of local ADI therapy was determined in xenopatients. Half of the GBM cell lines tested responded well toward ADI monotherapy. In those cell lines, viability decreased significantly (up to 50%). Responding cell lines were subjected to combination therapy experiments to test if any additive or even synergistic effects may be achieved. Such promising results were obtained in 2/3 cases. In cell lines HROG02, HROG05 and HROG10, ADI and Palomid 529 combinations were most effective yielding more than 70% killing after 2 rounds of treatment. Comparable boosted antitumoral effects were observed after adding chloroquine to ADI (>60% killing). Apoptosis, as well as cell cycle dysregulation were found to play a minor role. In some, but clearly not all cases, (epi-) genetic silencing of arginine synthesis pathway genes (argininosuccinate synthetase 1 and argininosuccinate lyase) explained obtained results. In vivo, ADI as well as the combination of ADI and SAHA efficiently controlled HROG05 xenograft growth, whereas adding Palomid 529 to ADI did not further increase the strong antitumoral effect of ADI. The cumulative in vitro and in vivo results proved ADI as a very promising candidate therapeutic, especially for development of adjuvant GBM combination treatments.

  10. Epidemiological study of erythromycin-resistant Streptococcus pyogenes from Korea and Japan by emm genotyping and multilocus sequence typing.

    PubMed

    Takahashi, Takashi; Arai, Kazuaki; Lee, Dong Hyun; Koh, Eun Ha; Yoshida, Haruno; Yano, Hisakazu; Kaku, Mitsuo; Kim, Sunjoo

    2016-01-01

    We determined the epidemiological characteristics of erythromycin (EM)-resistant Streptococcus pyogenes (group A streptococci, GAS) strains isolated from Korea and Japan, using emm genotyping and multilocus sequence typing (MLST). Clinical isolates of GAS had been collected from 1992 to 2012 in Korea and from 2004 to 2009 in Japan. EM resistance was determined by the microdilution method, and resistance genotypes were assessed by PCR. The emm genotyping and MLST were performed by DNA sequencing. The emm genotypes and sequence types (STs) were concordant in 143 (85.1%) of 168 EM-resistant GAS strains from Korea. ST36/emm12 (35.1%), ST52/emm28 (22.6%), and ST49/emm75 (16.1%) were the most common types. Most of the ST36 (93.9%) and ST52 (95.8%) strains harbored erm(B), whereas strains ST49, ST42, and ST15 contained mef(A). The concordance between emm genotypes and STs was 41 (93.2%) among 44 EM-resistant GAS strains from Japan. ST36/emm12 (34.1%), ST49/emm75 (18.2%), and ST28/emm1 (15.9%) were the major types. ST36 isolates harbored either erm(B) (56.3%) or mef(A) (37.5%), whereas isolates ST28, ST49, and ST38 carried only mef(A). The proportion of erm(B) and mef(A) was 66.1% and 33.3% in Korea and 22.7% and 68.2% in Japan, respectively. The common STs in Korea and Japan were ST36 and ST49, whereas ST52 was present only in Korea and ST28 only in Japan. Genotype erm(B) was predominant in Korea, whereas mef(A) was frequent in Japan. There were differences between Korea and Japan regarding the frequencies of emm genotypes, STs, and EM resistance genes among the EM-resistant GAS.

  11. Differences between Macrolide-Resistant and -Susceptible Streptococcus pyogenes: Importance of Clonal Properties in Addition to Antibiotic Consumption

    PubMed Central

    Silva-Costa, C.; Friães, A.; Melo-Cristino, J.

    2012-01-01

    A steady decline in macrolide resistance among Streptococcus pyogenes (group A streptococci [GAS]) in Portugal was reported during 1999 to 2006. This was accompanied by alterations in the prevalence of macrolide resistance phenotypes and in the clonal composition of the population. In order to test whether changes in the macrolide-resistant population reflected the same changing patterns of the overall population, we characterized both macrolide-susceptible and -resistant GAS associated with a diagnosis of tonsillo-pharyngitis recovered in the period from 2000 to 2005 in Portugal. Pulsed-field gel electrophoresis (PFGE) profiling was the best predictor of emm type and the only typing method that could discriminate clones associated with macrolide resistance and susceptibility within each emm type. Six PFGE clusters were significantly associated with macrolide susceptibility: T3-emm3-ST406, T4-emm4-ST39, T1-emm1-ST28, T6-emm6-ST382, B3264-emm89-ST101/ST408, and T2-emm2-ST55. Four PFGE clusters were associated with macrolide resistance: T4-emm4-ST39, T28-emm28-ST52, T12-emm22-ST46, and T1-emm1-ST28. We found no evidence for frequent ongoing horizontal transfer of macrolide resistance determinants. The diversity of the macrolide-resistant population was lower than that of susceptible isolates. The differences found between the two populations suggest that the macrolide-resistant population of GAS has its own dynamics, independent of the behavior of the susceptible population. PMID:22908153

  12. The Influence of Programmed Cell Death in Myeloid Cells on Host Resilience to Infection with Legionella pneumophila or Streptococcus pyogenes

    PubMed Central

    Gamradt, Pia; Xu, Yun; Gratz, Nina; Duncan, Kellyanne; Kobzik, Lester; Högler, Sandra; Decker, Thomas

    2016-01-01

    Pathogen clearance and host resilience/tolerance to infection are both important factors in surviving an infection. Cells of the myeloid lineage play important roles in both of these processes. Neutrophils, monocytes, macrophages, and dendritic cells all have important roles in initiation of the immune response and clearance of bacterial pathogens. If these cells are not properly regulated they can result in excessive inflammation and immunopathology leading to decreased host resilience. Programmed cell death (PCD) is one possible mechanism that myeloid cells may use to prevent excessive inflammation. Myeloid cell subsets play roles in tissue repair, immune response resolution, and maintenance of homeostasis, so excessive PCD may also influence host resilience in this way. In addition, myeloid cell death is one mechanism used to control pathogen replication and dissemination. Many of these functions for PCD have been well defined in vitro, but the role in vivo is less well understood. We created a mouse that constitutively expresses the pro-survival B-cell lymphoma (bcl)-2 protein in myeloid cells (CD68(bcl2tg), thus decreasing PCD specifically in myeloid cells. Using this mouse model we explored the impact that decreased cell death of these cells has on infection with two different bacterial pathogens, Legionella pneumophila and Streptococcus pyogenes. Both of these pathogens target multiple cell death pathways in myeloid cells, and the expression of bcl2 resulted in decreased PCD after infection. We examined both pathogen clearance and host resilience and found that myeloid cell death was crucial for host resilience. Surprisingly, the decreased myeloid PCD had minimal impact on pathogen clearance. These data indicate that the most important role of PCD during infection with these bacteria is to minimize inflammation and increase host resilience, not to aid in the clearance or prevent the spread of the pathogen. PMID:27973535

  13. Antibacterial activity and dual mechanisms of peptide analog derived from cell-penetrating peptide against Salmonella typhimurium and Streptococcus pyogenes.

    PubMed

    Li, Lirong; Shi, Yonghui; Cheserek, Maureen Jepkorir; Su, Guanfang; Le, Guowei

    2013-02-01

    A number of research have proven that antimicrobial peptides are of greatest potential as a new class of antibiotics. Antimicrobial peptides and cell-penetrating peptides share some similar structure characteristics. In our study, a new peptide analog, APP (GLARALTRLLRQLTRQLTRA) from the cell-penetrating peptide ppTG20 (GLFRALLRLLRSLWRLLLRA), was identified simultaneously with the antibacterial mechanism of APP against Salmonella typhimurium and Streptococcus pyogenes. APP displayed potent antibacterial activity against Gram-negative and Gram-positive strains. The minimum inhibitory concentration was in the range of 2 to 4 μM. APP displayed higher cell selectivity (about 42-fold increase) as compared to the parent peptide for it decreased hemolytic activity and increased antimicrobial activity. The calcein leakage from egg yolk L-α-phosphatidylcholine (EYPC)/egg yolk L-α-phosphatidyl-DL-glycerol and EYPC/cholesterol vesicles demonstrated that APP exhibited high selectivity. The antibacterial mechanism analysis indicated that APP induced membrane permeabilization in a kinetic manner for membrane lesions allowing O-nitrophenyl-β-D-galactoside uptake into cells and potassium release from APP-treated cells. Flow cytometry analysis demonstrated that APP induced bacterial live cell membrane damage. Circular dichroism, fluorescence spectra, and gel retardation analysis confirmed that APP interacted with DNA and intercalated into the DNA base pairs after penetrating the cell membrane. Cell cycle assay showed that APP affected DNA synthesis in the cell. Our results suggested that peptides derived from the cell-penetrating peptide have the potential for antimicrobial agent development, and APP exerts its antibacterial activity by damaging bacterial cell membranes and binding to bacterial DNA to inhibit cellular functions, ultimately leading to cell death.

  14. Unique Genomic Arrangements in an Invasive Serotype M23 Strain of Streptococcus pyogenes Identify Genes That Induce Hypervirulence

    PubMed Central

    Bao, Yunjuan; Liang, Zhong; Booyjzsen, Claire; Mayfield, Jeffrey A.; Li, Yang; Lee, Shaun W.; Ploplis, Victoria A.; Song, Hui

    2014-01-01

    The first genome sequence of a group A Streptococcus pyogenes serotype M23 (emm23) strain (M23ND), isolated from an invasive human infection, has been completed. The genome of this opacity factor-negative (SOF−) strain is composed of a circular chromosome of 1,846,477 bp. Gene profiling showed that this strain contained six phage-encoded and 24 chromosomally inherited well-known virulence factors, as well as 11 pseudogenes. The bacterium has acquired four large prophage elements, ΦM23ND.1 to ΦM23ND.4, harboring genes encoding streptococcal superantigen (ssa), streptococcal pyrogenic exotoxins (speC, speH, and speI), and DNases (spd1 and spd3), with phage integrase genes being present at one flank of each phage insertion, suggesting that the phages were integrated by horizontal gene transfer. Comparative analyses revealed unique large-scale genomic rearrangements that result in genomic rearrangements that differ from those of previously sequenced GAS strains. These rearrangements resulted in an imbalanced genomic architecture and translocations of chromosomal virulence genes. The covS sensor in M23ND was identified as a pseudogene, resulting in the attenuation of speB function and increased expression of the genes for the chromosomal virulence factors multiple-gene activator (mga), M protein (emm23), C5a peptidase (scpA), fibronectin-binding proteins (sfbI and fbp54), streptolysin O (slo), hyaluronic acid capsule (hasA), streptokinase (ska), and DNases (spd and spd3), which were verified by PCR. These genes are responsible for facilitating host epithelial cell binding and and/or immune evasion, thus further contributing to the virulence of M23ND. In conclusion, strain M23ND has become highly pathogenic as the result of a combination of multiple genetic factors, particularly gene composition and mutations, prophage integrations, unique genomic rearrangements, and regulated expression of critical virulence factors. PMID:25225265

  15. THE THERAPEUTIC ACTIVITY OF PENICILLINS F, G, K, AND X IN EXPERIMENTAL INFECTIONS WITH PNEUMOCOCCUS TYPE I AND STREPTOCOCCUS PYOGENES

    PubMed Central

    Eagle, Harry

    1947-01-01

    1. The relative bactericidal activities of penicillins F, G, K, and X against Type I pneumococcus in vitro were 60, 100, 180, and 135. The corresponding activities against Streptococcus pyogenes, strain C-203, were 75, 100, 115, and 145, respectively. 2. The total curative doses (CD50) of penicillins F, G, K, and X in pneumococcal infections of white mice (ten injections at 3 hour intervals) were 4.6, 3.8, 20, and 2.4 mg. per kg., respectively, or relative activities of 83, 100, 19, and 160, referred to G as 100. 3. The corresponding curative doses in streptococcal infections of white mice were 2.6, 1.3, 14.0, and 0.5 mg. per kg., or relative activities of 50, 100, 9, and 260. 4. Penicillin K was therefore one-tenth as active in vivo as would be implied by its bactericidal activity in vitro. This probably reflects its rapid inactivation in vivo, evidenced by the low and evanescent blood levels observed in both rabbits and man, and the low urinary recovery of this species of penicillin. 5. Penicillin X was significantly more active therapeutically than its bactericidal activity in vitro would imply. This probably reflects its slower inactivation in vivo, evidenced by the somewhat higher and more prolonged blood levels afforded by this penicillin in comparison with penicillin G. Judged by the mouse infections with the strains here used, penicillin X is the penicillin of choice in the treatment of infections with pneumococcus Type I and hemolytic streptococci. 6. The curative dose of penicillin in streptococcal and pneumococcal infections paralleled the varying susceptibility of these organisms to penicillin in vitro. PMID:19871606

  16. Identification of Residues Responsible for the Defective Virulence Gene Regulator Mga Produced by a Natural Mutant of Streptococcus pyogenes

    PubMed Central

    Vahling, Cheryl M.; McIver, Kevin S.

    2005-01-01

    Mga is a transcriptional regulator in the pathogen Streptococcus pyogenes that positively activates several important virulence genes involved in colonization and immune evasion in the human host. A naturally occurring mutant of Mga that is defective in its ability to activate transcription has been identified in the serotype M50 strain B514-Sm. Sequence alignment of the defective M50 Mga with the fully functional Mga from serotypes M4 and M49 revealed only three amino acid changes that might result in a defective protein. Electrophoretic mobility shift assays using purified M50 and M4 maltose binding protein-Mga found that both exhibited DNA-binding activity towards regulated promoters. Thus, the significance of each residue for the functionality of M50 Mga was explored through introduction of “gain-of-function” mutations based on M4 Mga. Transcriptional studies of the mutant alleles under both constitutive (PrpsL) and autoactivated (Pmga4) promoters illustrated that an arginine-to-methionine change at position 461 of M50 Mga protein fully restored activation of downstream genes. Western blot analyses of steady-state Mga levels suggest that the M461 residue may play a role in overall conformation and protein stability of Mga. However, despite the conservation of the M461 protein among all other Mga proteins, it does not appear to be necessary for activity in a divergent M6 Mga. These studies highlight the potential differences that exist between divergent Mga proteins in this important human pathogen. PMID:16109937

  17. CcpA and LacD.1 Affect Temporal Regulation of Streptococcus pyogenes Virulence Genes ▿ †

    PubMed Central

    Kietzman, Colin C.; Caparon, Michael G.

    2010-01-01

    Production of H2O2 follows a growth phase-dependent pattern that mimics that of many virulence factors of Streptococcus pyogenes. To gain greater insight into mechanisms coupling virulence factor expression to growth phase, we investigated the molecular basis for H2O2 generation and its regulation. Deletion of the gene encoding lactate oxidase (lctO) or culture in the presence of glucose eliminated H2O2 production, implicating carbohydrate regulation of lctO as a key element of growth phase control. In examining known carbohydrate-responsive regulators, deletion of the gene encoding CcpA but not that encoding LacD.1 resulted in both derepression and an uncoupling of lctO transcription from its growth phase pattern. Expanding this analysis to additional virulence factors demonstrated both negative (cfa, encoding CAMP factor) and positive (speB, encoding a cysteine protease) regulation by CcpA and that CcpA mutants were highly cytotoxic for cultured macrophages. This latter property resulted from enhanced transcription of the streptolysin S biogenesis operon. Examination of CcpA-promoter interactions using a DNA pull-down assay mimicking physiological conditions showed direct binding to the promoters of lctO and speB but not those of sagA. CcpA but not LacD.1 mutants were attenuated in a murine model of soft-tissue infection, and analysis of gene expression in infected tissue indicated that CcpA mutants had altered expression of lctO, cfa, and speB but not the indirectly regulated sagA gene. Taken together, these data show that CcpA regulates virulence genes via at least three distinct mechanisms and that disruption of growth phase regulation alters transcriptional patterns in infected tissues. PMID:19841076

  18. Dissecting a bacterial collagen domain from Streptococcus pyogenes: sequence and length-dependent variations in triple helix stability and folding.

    PubMed

    Yu, Zhuoxin; Brodsky, Barbara; Inouye, Masayori

    2011-05-27

    To better investigate the relationship between sequence, stability, and folding, the Streptococcus pyogenes collagenous domain CL (Gly-Xaa-Yaa)(79) was divided to create three recombinant triple helix subdomains A, B, and C of almost equal size with distinctive amino acid features: an A domain high in polar residues, a B domain containing the highest concentration of Pro residues, and a very highly charged C domain. Each segment was expressed as a monomer, a linear dimer, and a linear trimer fused with the trimerization domain (V domain) in Escherichia coli. All recombinant proteins studied formed stable triple helical structures, but the stability varied depending on the amino acid sequence in the A, B, and C segments and increased as the triple helix got longer. V-AAA was found to melt at a much lower temperature (31.0 °C) than V-ABC (V-CL), whereas V-BBB melted at almost the same temperature (∼36-37 °C). When heat-denatured, the V domain enhanced refolding for all of the constructs; however, the folding rate was affected by their amino acid sequences and became reduced for longer constructs. The folding rates of all the other constructs were lower than that of the natural V-ABC protein. Amino acid substitution mutations at all Pro residues in the C fragment dramatically decreased stability but increased the folding rate. These results indicate that the thermostability of the bacterial collagen is dominated by the most stable domain in the same manner as found with eukaryotic collagens.

  19. Copper Tolerance and Characterization of a Copper-Responsive Operon, copYAZ, in an M1T1 Clinical Strain of Streptococcus pyogenes

    PubMed Central

    Gordon, Lily D.; Fang, Zhong; Holder, Robert C.; Reid, Sean D.

    2015-01-01

    ABSTRACT Infection with Streptococcus pyogenes is associated with a breadth of clinical manifestations ranging from mild pharyngitis to severe necrotizing fasciitis. Elevated levels of intracellular copper are highly toxic to this bacterium, and thus, the microbe must tightly regulate the level of this metal ion by one or more mechanisms, which have, to date, not been clearly defined. In this study, we have identified two virulence mechanisms by which S. pyogenes protects itself against copper toxicity. We defined a set of putative genes, copY (for a regulator), copA (for a P1-type ATPase), and copZ (for a copper chaperone), whose expression is regulated by copper. Our results indicate that these genes are highly conserved among a range of clinical S. pyogenes isolates. The copY, copA, and copZ genes are induced by copper and are transcribed as a single unit. Heterologous expression assays revealed that S. pyogenes CopA can confer copper tolerance in a copper-sensitive Escherichia coli mutant by preventing the accumulation of toxic levels of copper, a finding that is consistent with a role for CopA in copper export. Evaluation of the effect of copper stress on S. pyogenes in a planktonic or biofilm state revealed that biofilms may aid in protection during initial exposure to copper. However, copper stress appears to prevent the shift from the planktonic to the biofilm state. Therefore, our results indicate that S. pyogenes may use several virulence mechanisms, including altered gene expression and a transition to and from planktonic and biofilm states, to promote survival during copper stress. IMPORTANCE Bacterial pathogens encounter multiple stressors at the host-pathogen interface. This study evaluates a virulence mechanism(s) utilized by S. pyogenes to combat copper at sites of infection. A better understanding of pathogen tolerance to stressors such as copper is necessary to determine how host-pathogen interactions impact bacterial survival during infections

  20. Paediatric obstructive sleep apnoea syndrome (OSAS) is associated with tonsil colonisation by Streptococcus pyogenes

    PubMed Central

    Viciani, Elisa; Montagnani, Francesca; Tavarini, Simona; Tordini, Giacinta; Maccari, Silvia; Morandi, Matteo; Faenzi, Elisa; Biagini, Cesare; Romano, Antonio; Salerni, Lorenzo; Finco, Oretta; Lazzi, Stefano; Ruggiero, Paolo; De Luca, Andrea; Barocchi, Michèle A.; Manetti, Andrea G. O.

    2016-01-01

    The involvement of pathogenic bacteria in obstructive sleep apnoea syndrome (OSAS) has yet to be elucidated. We investigated the possible role of group A streptococcus (GAS) in OSAS pathogenesis. In 40 tonsillectomized patients affected by OSAS and 80 healthy controls, significant (p < 0.0001) association of GAS with paediatric OSAS was found. Supernatant from streptolysin O (SLO)-producing GAS induced production of cysteinyl leukotrienes (CysLTs) in tonsil mononuclear cells (TMCs). CysLTs-treated TMCs showed significant (p < 0.05) proliferation of CD4+ T, CD19+ and CD19+CD27+CD38+ B lymphocytes. We discovered a SLO-dependent activation of CysLTs production through a pathway involving TOLL-like receptor 4 (TLR4), TIR-domain-containing adapter-inducing interferon-β (TRIF), Myeloid differentiation primary response gene 88 (MyD88), and p38 MAP Kinase. In conclusion, we hypothesise that GAS may contribute to paediatric tonsillar hyperplasia through CysLTs production induced by SLO, and this might explain its association with OSAS. PMID:26860261

  1. Dermal mast cells reduce progressive tissue necrosis caused by subcutaneous infection with Streptococcus pyogenes in mice.

    PubMed

    Matsui, Hidenori; Sekiya, Yukie; Takahashi, Tetsufumi; Nakamura, Masahiko; Imanishi, Ken'ichi; Yoshida, Haruno; Murayama, Somay Yamagata; Takahashi, Takashi; Tsuchimoto, Kanji; Uchiyama, Takehiko; Ubukata, Kimiko

    2011-01-01

    A single subcutaneous (s.c.) infection with 1×10(7) c.f.u. GAS472, a group A streptococcus (GAS) serotype M1 strain isolated from the blood of a patient suffering from streptococcal toxic shock syndrome, led to severe damage of striated muscle layers in the feet of mast cell (MC)-deficient WBB6F(1)-Kit(W)/Kit(W-v) (W/W(v)) mice 72 h after infection. In contrast, no damage was recognized in striated muscle layers in the feet of the control WBB6F(1)-Kit(+/+) (+/+) mice 72 h after infection. In addition, adoptively transferred MCs reduced progressive tissue necrosis of the feet of W/W(v) mice after infection. However, there was no significant difference in the mortality rates between the W/W(v) and +/+ mice, or between the human CD46-expressing transgenic (Tg) mouse bone marrow-derived cultured MC-reconstituted W/W(v) and non-Tg mouse bone marrow-derived cultured MC-reconstituted W/W(v) mice after infection. Consequently, although MCs can help to reduce the severity of necrosis of the feet caused by s.c. infection with GAS472, such reduction of tissue necrosis scarcely improves the mortality rates of these mice. Moreover, human CD46 does not play a crucial role in the MC-mediated innate immune defence against GAS infection.

  2. Identification and Characterization of a Novel Heme-Associated Cell Surface Protein Made by Streptococcus pyogenes

    PubMed Central

    Lei, Benfang; Smoot, Laura M.; Menning, Heather M.; Voyich, Jovanka M.; Kala, Subbarao V.; Deleo, Frank R.; Reid, Sean D.; Musser, James M.

    2002-01-01

    Analysis of the genome sequence of a serotype M1 group A Streptococcus (GAS) strain identified a gene encoding a previously undescribed putative cell surface protein. The gene was cloned from a serotype M1 strain, and the recombinant protein was overexpressed in Escherichia coli and purified to homogeneity. The purified protein was associated with heme in a 1:1 stoichiometry. This streptococcal heme-associated protein, designated Shp, was produced in vitro by GAS, located on the bacterial cell surface, and accessible to specific antibody raised against the purified recombinant protein. Mice inoculated subcutaneously with GAS and humans with invasive infections and pharyngitis caused by GAS seroconverted to Shp, indicating that Shp was produced in vivo. The blood of mice actively immunized with Shp had significantly higher bactericidal activity than the blood of unimmunized mice. The shp gene was cotranscribed with eight contiguous genes, including homologues of an ABC transporter involved in iron uptake in gram-negative bacteria. Our results indicate that Shp is a novel cell surface heme-associated protein. PMID:12117961

  3. Structural and functional analysis of RopB: A major virulence regulator in Streptococcus pyogenes

    DOE PAGES

    Makthal, Nishanth; Gavagan, Maire; Do, Hackwon; ...

    2016-02-19

    Group A Streptococcus (GAS) is an exclusive human pathogen that causes significant disease burden. Global regulator RopB of GAS controls the expression of several major virulence factors including secreted protease SpeB during high cell density. However, the molecular mechanism for RopB-dependent speB expression remains unclear. To understand the mechanism of transcription activation by RopB, we determined the crystal structure of the C-terminal domain of RopB. RopB-CTD has the TPR motif, a signature motif involved in protein-peptide interactions and shares significant structural homology with the quorum sensing RRNPP family regulators. Characterization of the high cell density-specific cell-free growth medium demonstrated themore » presence of a low molecular weight proteinaceous secreted factor that upregulates RopB-dependent speB expression. Together, these results suggest that RopB and its cognate peptide signals constitute an intercellular signalling machinery that controls the virulence gene expression in concert with population density. Structure-guided mutational analyses of RopB dimer interface demonstrated that single alanine substitutions at this critical interface significantly altered RopB-dependent speB expression and attenuated GAS virulence. Finally, results presented here suggested that a properly aligned RopB dimer interface is important for GAS pathogenesis and highlighted the dimerization interactions as a plausible therapeutic target for the development of novel antimicrobials.« less

  4. Molecular analysis of the role of streptococcal pyrogenic Exotoxin A (SPEA) in invasive soft-tissue infection resulting from Streptococcus pyogenes.

    PubMed

    Sriskandan, S; Unnikrishnan, M; Krausz, T; Cohen, J

    1999-08-01

    Epidemiological studies strongly implicate the bacterial superantigen, streptococcal pyrogenic exotoxin A (SPEA), in the pathogenesis of necrotizing soft-tissue infection and toxic shock syndrome resulting from Streptococcus pyogenes. SPEA can act as a superantigen and cellular toxin ex vivo, but its role during invasive streptococcal infection is unclear. We have disrupted the wild-type spea gene in an M1 streptococcal isolate. Supernatants from toxin-negative mutant bacteria demonstrated a 50% reduction in pro-mitogenic activity in HLA DQ-positive murine splenocyte culture, and up to 20% reduction in activity in human PBMC culture. Mutant and wild-type bacteria were then compared in mouse models of bacteraemia and streptococcal muscle infection. Disruption of spea was not associated with attenuation of virulence in either model. Indeed, a paradoxical increase in mutant strain-induced mortality was seen after intravenous infection. Intramuscular infection with the SPEA-negative mutant led to increased bacteraemia at 24 h and a reduction in neutrophils at the site of primary muscle infection. Purified SPEA led to a dose-dependent increase in peritoneal neutrophils 6 h after administration. SPEA is not a critical virulence factor in invasive soft-tissue infection or bacteraemia caused by S. pyogenes, and it could have a protective role in murine immunity to pyogenic infection. The role of this toxin may be different in hosts with augmented superantigen responsiveness.

  5. Surface Export of GAPDH/SDH, a Glycolytic Enzyme, Is Essential for Streptococcus pyogenes Virulence

    PubMed Central

    Jin, Hong; Agarwal, Shivangi; Agarwal, Shivani; Pancholi, Vijay

    2011-01-01

    ABSTRACT Streptococcal surface dehydrogenase (SDH) (glyceraldehyde-3-phosphate dehydrogenase [GAPDH]) is an anchorless major multifunctional surface protein in group A Streptococcus (GAS) with the ability to bind important mammalian proteins, including plasmin(ogen). Although several biological properties of SDH are suggestive of its possible role in GAS virulence, its direct role in GAS pathogenesis has not been ascertained because it is essential for GAS survival. Thus, it has remained enigmatic as to “how and why” SDH/GAPDH is exported onto the bacterial surface. The present investigation highlights “why” SDH is exported onto the GAS surface. Differential microarray-based genome-wide transcript abundance analysis was carried out using a specific mutant, which was created by inserting a hydrophobic tail at the C-terminal end of SDH (M1-SDHHBtail) and thus preventing its exportation onto the GAS surface. This analysis revealed downregulation of the majority of genes involved in GAS virulence and genes belonging to carbohydrate and amino acid metabolism and upregulation of those related to lipid metabolism. The complete attenuation of this mutant for virulence in the mouse model and the decreased and increased virulence of the wild-type and mutant strains postcomplementation with SDHHBtail and SDH, respectively, indicated that the SDH surface export indeed regulates GAS virulence. M1-SDHHBtail also displayed unaltered growth patterns, increased intracellular ATP concentration and Hpr double phosphorylation, and significantly reduced pH tolerance, streptolysin S, and SpeB activities. These phenotypic and physiological changes observed in the mutant despite the unaltered expression levels of established transcriptional regulators further highlight the fact that SDH interfaces with many regulators and its surface exportation is essential for GAS virulence. PMID:21628503

  6. CD46 transgenic mouse model of necrotizing fasciitis caused by Streptococcus pyogenes infection.

    PubMed

    Matsui, Hidenori; Sekiya, Yukie; Nakamura, Masahiko; Murayama, Somay Yamagata; Yoshida, Haruno; Takahashi, Tetsufumi; Imanishi, Ken'ichi; Tsuchimoto, Kanji; Uchiyama, Takehiko; Sunakawa, Keisuke; Ubukata, Kimiko

    2009-11-01

    We developed a human CD46-expressing transgenic (Tg) mouse model of subcutaneous (s.c.) infection into both hind footpads with clinically isolated 11 group A streptococcus (GAS) serotype M1 strains. When the severity levels of foot lesions at 72 h and the mortality rates by 336 h were compared after s.c. infection with 1x10(7) CFU of each GAS strain, the GAS472 strain, isolated from the blood of a patient suffering from streptococcal toxic shock syndrome (STSS), induced the highest severity levels and mortality rates. GAS472 led to a 100% mortality rate in CD46 Tg mice after only 168 h postinfection through the supervention of severe necrotizing fasciitis (NF) of the feet. In contrast, GAS472 led to a 10% mortality rate in non-Tg mice through the supervention of partial necrotizing cutaneous lesions of the feet. The footpad skin sections of CD46 Tg mice showed hemorrhaging and necrotic striated muscle layers in the dermis, along with the exfoliation of epidermis with intracellular edema until 48 h after s.c. infection with GAS472. Thereafter, the bacteria proliferated, reaching a 90-fold or 7-fold increase in the livers of CD46 Tg mice or non-Tg mice, respectively, for 24 h between 48 and 72 h after s.c. infection with GAS472. As a result, the infected CD46 Tg mice appeared to suffer severe liver injuries. These findings suggest that human CD46 enhanced the progression of NF in the feet and the exponential growth of bacteria in deep tissues, leading to death.

  7. Revelation of susceptibility differences due to Hg(II) accumulation in Streptococcus pyogenes against CX-AgNPs and Cefixime by atomic force microscopy.

    PubMed

    Rasheed, Wasia; Shah, Muhammad Raza; Perveen, Samina; Ahmed, Shakil; Uzzaman, Sami

    2017-08-16

    Solution based method for the formation of chemically modified silver nanoparticles (CX-AgNPs) using Cefixime as stabilizing and reducing agent was developed. The CX-AgNPs were characterized by AFM, UV-visible, FT-IR and MALDI-TOF MS. Bactericidal efficiency of CX-AgNPs and Cefixime against Streptococcus pyogenes was evaluated. Afterwards, susceptibility differences of Streptococcus pyogenes due to accumulation of Hg(II) against CX-AgNPs and Cefixime were estimated and validated through Atomic force microscopy. Selectivity and sensitivity of CX-AgNPs against Hg(II) was evaluated in a systematic manner. The CX-AgNPs was titrated against optically silent Hg(II) which induced enhancement in the SPR band of CX-AgNPs. The increase in intensity of SPR band of CX-AgNPs was determined to be proportionate to the concentration of Hg(II) in the range of 33.3-700µM obeying linear regression equation of y = 0.125x + 8.962 with the detection limit of 0.10µM and the coefficient of determination equals to 0.985 (n = 3). The association constant Ka of CX-AgNPs-Hg(II) was found to be 386.0095mol(-1)dm(3) by using the Benesi Hildebrand plot. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Mry, a trans-acting positive regulator of the M protein gene of Streptococcus pyogenes with similarity to the receptor proteins of two-component regulatory systems.

    PubMed Central

    Perez-Casal, J; Caparon, M G; Scott, J R

    1991-01-01

    In the Streptococcus pyogenes M6 strain D471, an insertion of the conjugative transposon Tn916 into a region 2 kb upstream of the promoter of emm6 (the structural gene for the M protein) rendered the strain M negative (M. G. Caparon and J. R. Scott, Proc. Natl. Acad. Sci. USA 84:8677-8681, 1987). In the present work, we show that this insertion mutation, mry-1, is 244 bp upstream of an open reading frame encoding a protein we call Mry. This protein is visible on a gel after transcription and translation in vitro. We have developed a technique for complementation analysis in S. pyogenes and have used it to show that the wild-type mry gene is dominant to two mutant alleles. This dominance indicates that Mry acts in trans as a positive regulator of the emm6 gene. The translated DNA sequence of mry has two regions of similarity to the motif common to the receptor protein of two-component regulatory systems. In addition, the N terminus of Mry has two regions resembling a helix-turn-helix motif. Mry does not appear to be a global regulator of virulence determinants in the group A streptococcus because there is no effect of the mry-1 mutation on production of the hyaluronic acid capsule or streptokinase. Images PMID:1849511

  9. Analysis of the role of CovR and CovS in the dissemination of Streptococcus pyogenes in invasive skin disease.

    PubMed

    Dalton, Tracy L; Hobb, Rhonda I; Scott, June R

    2006-05-01

    The global regulatory two-component system CovR/S controls expression of about 15% of the Streptococcus pyogenes (group A streptococcus; GAS) genome. Recently, we found that CovS plays a pivotal role in general stress response of this strictly human pathogen. Therefore, we expected that both CovS and CovR might affect virulence. In this work, mice were inoculated subcutaneously with isogenic nonpolar covR and covS deletion-substitution mutants and the isogenic wild-type strain. The covS mutant behaved like the wild-type parental strain in terms of resulting lesion appearance and invasive disease leading to death. This is in agreement with previous results suggesting that the absence of its cognate sensor kinase does not affect the ability of CovR to become phosphorylated and cause repression of its regulon. However, two different covR deletion-substitution mutants caused significantly less invasive disease and death in the mice than the wild-type parental strain, although the local lesions produced by the covR mutants were more severe and purulent than those resulting from the wild-type GAS strain. Thus, it appears that production of CovR increases the ability of S. pyogenes to cause severe invasive disease in this mouse model and therefore is an important virulence factor for this organism.

  10. Comparative activity of garenoxacin and other agents by susceptibility and time-kill testing against Staphylococcus aureus, Streptococcus pyogenes and respiratory pathogens.

    PubMed

    Noviello, Silvana; Ianniello, Filomena; Leone, Sebastiano; Esposito, Silvano

    2003-11-01

    Garenoxacin is a novel des-F(6)quinolone that has shown excellent antimicrobial activity against a wide range of clinically important microorganisms. In this study, its activity was examined, in comparison with that of other antimicrobial agents, by susceptibility and time-kill testing against Staphylococcus aureus, Streptococcus pyogenes and respiratory pathogens. Overall, 200 bacterial strains were tested. The antimicrobial activity of garenoxacin was compared with that of ciprofloxacin, levofloxacin, moxifloxacin, amoxicillin, co-amoxiclav, cefuroxime, cefotaxime, ceftriaxone, imipenem, erythromycin and clarithromycin. In addition, the bactericidal activity of garenoxacin, moxifloxacin, levofloxacin and ciprofloxacin was evaluated by time-kill analysis against four strains each of staphylococci [two methicillin-susceptible (MSSA) and two methicillin-resistant (MRSA)], pneumococci (two penicillin-susceptible and two penicillin-resistant) and Streptococcus pyogenes (two erythromycin-susceptible and two erythromycin-resistant). Antibiotics were tested at concentrations 1-8 x MIC. MIC90 values of garenoxacin for the MSSA and MRSA strains were 0.03 and 2 mg/L, respectively. Among all the quinolones tested, garenoxacin yielded the lowest MIC values against all pneumococci (MIC90 0.12 mg/L) irrespective of macrolide resistance; the rank order of activity was garenoxacin> moxifloxacin>levofloxacin>ciprofloxacin. Excellent activity was shown also against Haemophilus influenzae (MIC90 pyogenes were inhibited at garenoxacin concentrations equal to 0.25 mg/L, its activity not being influenced by macrolide susceptibility. Garenoxacin was rapidly bactericidal against staphylococci, producing a >or= 3 log10 decrease in viable counts (cfu/mL) within 3 h at 4 x MIC, whereas a moderate, slower killing rate was observed versus streptococci. This investigational des-F(6)quinolone represents a

  11. Activity of retapamulin against Streptococcus pyogenes and Staphylococcus aureus evaluated by agar dilution, microdilution, E-test, and disk diffusion methodologies.

    PubMed

    Pankuch, Glenn A; Lin, Gengrong; Hoellman, Dianne B; Good, Caryn E; Jacobs, Michael R; Appelbaum, Peter C

    2006-05-01

    The in vitro activity of retapamulin against 106 Staphylococcus aureus isolates and 109 Streptococcus pyogenes isolates was evaluated by the agar dilution, broth microdilution, E-test, and disk diffusion methodologies. Where possible, the tests were performed by using the CLSI methodology. The results of agar dilution, broth microdilution, and E-test (all with incubation in ambient air) for S. aureus yielded similar MICs, in the range of 0.03 to 0.25 microg/ml. These values corresponded to zone diameters between 25 and 33 mm by the use of a 2-microg retapamulin disk. Overall, 99% of the agar dilution results and 95% of E-test results for S. aureus were within +/-1 dilution of the microdilution results. For S. pyogenes, the MICs obtained by the agar and broth microdilution methods (both after incubation in ambient air) were in the range of 0.008 to 0.03 microg/ml, and E-test MICs (with incubation in ambient air) were 0.016 to 0.06 microg/ml. For S. pyogenes, 100% of the agar dilution MIC results were within +/-1 dilution of the broth microdilution results. E-test MICs (after incubation in ambient air) were within +/-1 and +/-2 dilutions of the broth microdilution results for 76% and 99% of the isolates, respectively. E-test MICs for S. pyogenes strains in CO(2) were up to 4 dilutions higher than those in ambient air. Therefore, it is recommended that when retapamulin MICs are determined by E-test, incubation be done in ambient air and not in CO(2), due to the adverse effect of CO(2) on the activity of this compound. Diffusion zones (with incubation in CO(2)) for S. pyogenes were 18 to 24 mm. Retapamulin MICs for all strains by all methods (with incubation in ambient air) were < or =0.25 microg/ml. These results demonstrate that S. pyogenes (including macrolide-resistant strains) and S. aureus (including methicillin-resistant and vancomycin-nonsusceptible strains) are inhibited by very low concentrations of retapamulin and that all four testing methods are

  12. Activity of Retapamulin against Streptococcus pyogenes and Staphylococcus aureus Evaluated by Agar Dilution, Microdilution, E-Test, and Disk Diffusion Methodologies

    PubMed Central

    Pankuch, Glenn A.; Lin, Gengrong; Hoellman, Dianne B.; Good, Caryn E.; Jacobs, Michael R.; Appelbaum, Peter C.

    2006-01-01

    The in vitro activity of retapamulin against 106 Staphylococcus aureus isolates and 109 Streptococcus pyogenes isolates was evaluated by the agar dilution, broth microdilution, E-test, and disk diffusion methodologies. Where possible, the tests were performed by using the CLSI methodology. The results of agar dilution, broth microdilution, and E-test (all with incubation in ambient air) for S. aureus yielded similar MICs, in the range of 0.03 to 0.25 μg/ml. These values corresponded to zone diameters between 25 and 33 mm by the use of a 2-μg retapamulin disk. Overall, 99% of the agar dilution results and 95% of E-test results for S. aureus were within ±1 dilution of the microdilution results. For S. pyogenes, the MICs obtained by the agar and broth microdilution methods (both after incubation in ambient air) were in the range of 0.008 to 0.03 μg/ml, and E-test MICs (with incubation in ambient air) were 0.016 to 0.06 μg/ml. For S. pyogenes, 100% of the agar dilution MIC results were within ±1 dilution of the broth microdilution results. E-test MICs (after incubation in ambient air) were within ±1 and ±2 dilutions of the broth microdilution results for 76% and 99% of the isolates, respectively. E-test MICs for S. pyogenes strains in CO2 were up to 4 dilutions higher than those in ambient air. Therefore, it is recommended that when retapamulin MICs are determined by E-test, incubation be done in ambient air and not in CO2, due to the adverse effect of CO2 on the activity of this compound. Diffusion zones (with incubation in CO2) for S. pyogenes were 18 to 24 mm. Retapamulin MICs for all strains by all methods (with incubation in ambient air) were ≤0.25 μg/ml. These results demonstrate that S. pyogenes (including macrolide-resistant strains) and S. aureus (including methicillin-resistant and vancomycin-nonsusceptible strains) are inhibited by very low concentrations of retapamulin and that all four testing methods are satisfactory for use for susceptibility

  13. Application of an enzyme-labeled antigen method for visualizing plasma cells producing antibodies against Strep A, a carbohydrate antigen of Streptococcus pyogenes, in recurrent tonsillitis.

    PubMed

    Onouchi, Takanori; Mizutani, Yasuyoshi; Shiogama, Kazuya; Inada, Ken-ichi; Okada, Tatsuyoshi; Naito, Kensei; Tsutsumi, Yutaka

    2015-01-01

    Streptococcus pyogenes is the main causative pathogen of recurrent tonsillitis. Histologically, lesions of recurrent tonsillitis contain numerous plasma cells. Strep A is an antigenic carbohydrate molecule on the cell wall of S. pyogenes. As expected, plasma cells in subjects with recurrent tonsillitis secrete antibodies against Strep A. The enzyme-labeled antigen method is a novel histochemical technique that visualizes specific antibody-producing cells in tissue sections by employing a biotin-labeled antigen as a probe. The purpose of the present study was to visualize plasma cells producing antibodies reactive with Strep A in recurrent tonsillitis. Firstly, the lymph nodes of rats immunized with boiled S. pyogenes were paraformaldehyde-fixed and specific plasma cells localized in frozen sections with biotinylated Strep A. Secondly, an enzyme-labeled antigen method was used on human tonsil surgically removed from 12 patients with recurrent tonsillitis. S. pyogenes genomes were PCR-detected in all 12 specimens. The emm genotypes belonged to emm12 in nine specimens and emm1 in three. Plasma cells producing anti-Strep A antibodies were demonstrated in prefixed frozen sections of rat lymph nodes, 8/12 human specimens from patients with recurrent tonsillitis but not in two control tonsils. In human tonsils, Strep A-reactive plasma cells were observed within the reticular squamous mucosa and just below the mucosa, and the specific antibodies belonged to either IgA or IgG classes. Our technique is effective in visualizing immunocytes producing specific antibodies against the bacterial carbohydrate antigen, and is thus a novel histochemical tool for analyzing immune reactions in infectious disorders. © 2014 The Authors. Microbiology and Immunology Published by The Societies and Wiley Publishing Asia Pty Ltd.

  14. Virulence Gene Pool Detected in Bovine Group C Streptococcus dysgalactiae subsp. dysgalactiae Isolates by Use of a Group A S. pyogenes Virulence Microarray ▿

    PubMed Central

    Rato, Márcia G.; Nerlich, Andreas; Bergmann, René; Bexiga, Ricardo; Nunes, Sandro F.; Vilela, Cristina L.; Santos-Sanches, Ilda; Chhatwal, Gursharan S.

    2011-01-01

    A custom-designed microarray containing 220 virulence genes of Streptococcus pyogenes (group A Streptococcus [GAS]) was used to test group C Streptococcus dysgalactiae subsp. dysgalactiae (GCS) field strains causing bovine mastitis and group C or group G Streptococcus dysgalactiae subsp. equisimilis (GCS/GGS) isolates from human infections, with the latter being used for comparative purposes, for the presence of virulence genes. All bovine and all human isolates carried a fraction of the 220 genes (23% and 39%, respectively). The virulence genes encoding streptolysin S, glyceraldehyde-3-phosphate dehydrogenase, the plasminogen-binding M-like protein PAM, and the collagen-like protein SclB were detected in the majority of both bovine and human isolates (94 to 100%). Virulence factors, usually carried by human beta-hemolytic streptococcal pathogens, such as streptokinase, laminin-binding protein, and the C5a peptidase precursor, were detected in all human isolates but not in bovine isolates. Additionally, GAS bacteriophage-associated virulence genes encoding superantigens, DNase, and/or streptodornase were detected in bovine isolates (72%) but not in the human isolates. Determinants located in non-bacteriophage-related mobile elements, such as the gene encoding R28, were detected in all bovine and human isolates. Several virulence genes, including genes of bacteriophage origin, were shown to be expressed by reverse transcriptase PCR (RT-PCR). Phylogenetic analysis of superantigen gene sequences revealed a high level (>98%) of identity among genes of bovine GCS, of the horse pathogen Streptococcus equi subsp. equi, and of the human pathogen GAS. Our findings indicate that alpha-hemolytic bovine GCS, an important mastitis pathogen and considered to be a nonhuman pathogen, carries important virulence factors responsible for virulence and pathogenesis in humans. PMID:21525223

  15. In vitro pharmacokinetic/pharmacodynamic activity of NXL103 versus clindamycin and linezolid against clinical Staphylococcus aureus and Streptococcus pyogenes isolates.

    PubMed

    Vidaillac, Celine; Parra-Ruiz, Jorge; Winterfield, Patricia; Rybak, Michael J

    2011-10-01

    NXL103 (linopristin/flopristin, 30/70) is a novel oral streptogramin combination with activity against a large variety of multidrug-resistant Gram-positive pathogens. The objective of this study was to evaluate the in vitro activity of NXL103 in comparison with oral comparators (clindamycin and linezolid). Six clinical isolates [four meticillin-resistant Staphylococcus aureus (MRSA) and two Streptococcus pyogenes] were exposed for 48 h in an in vitro pharmacokinetic/pharmacodynamic (PK/PD) model at a starting inoculum of ca. 10(6) colony-forming units (CFU)/mL. Antimicrobial simulations included NXL103 500 mg every 12 h, linezolid 600 mg every 12 h and clindamycin 450 mg every 6 h. Bactericidal and static effects were defined as ≥3log(10) and <3log(10) CFU/mL kill from the starting inoculum, respectively. Experiments were performed in duplicate to ensure reproducibility, and differences between regimens were evaluated by analysis of variance (ANOVA) with Tukey's post-hoc test. NXL103 exhibited lower minimum inhibitory concentrations than comparators, with values ≤0.06 mg/L for S. pyogenes and 0.125-0.25 mg/L for MRSA isolates. In the PK/PD model, NXL103 demonstrated significantly better activity than linezolid and clindamycin (P<0.05), achieving sustained bactericidal activity within <2 h against S. pyogenes strains and between 7.3-32 h against MRSA isolates. In contrast, linezolid only exhibited a static effect, whereas clindamycin achieved 3log(10) kill at 6h against the unique clindamycin-susceptible S. pyogenes strain evaluated. In conclusion, at therapeutic concentrations NXL103 exhibits promising activity against both MRSA and S. pyogenes strains, including clindamycin-resistant organisms. Further in vitro and in vivo experiments are warranted to explore the therapeutic benefit of NXL103 for the treatment of Gram-positive skin and soft-tissue infections. Copyright © 2011 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

  16. Substitution of cysteine 192 in a highly conserved Streptococcus pyogenes extracellular cysteine protease (interleukin 1beta convertase) alters proteolytic activity and ablates zymogen processing.

    PubMed Central

    Musser, J M; Stockbauer, K; Kapur, V; Rudgers, G W

    1996-01-01

    Virtually all strains of the human pathogenic bacterium Streptococcus pyogenes express a highly conserved extracellular cysteine protease. The protein is made as an inactive zymogen of 40,000 Da and undergoes autocatalytic truncation to result in a 28,000-Da active protease. Numerous independent lines of investigation suggest that this enzyme participates in one or more phases of host-parasite interaction, such as inflammation and soft tissue invasion. Replacement of the single cysteine residue (C-192) with serine (C192S mutation) resulted in loss of detectable proteolytic activity against bovine casein, human fibronectin, and the low-molecular-weight synthetic substrate 7-amino-4-trifluoromethyl coumarin. The C192S mutant molecule does not undergo autocatalytic processing of zymogen to mature form. Taken together, these data support the hypothesis that C-192 participates in active-site formation and enzyme catalysis. PMID:8675287

  17. Substitution of cysteine 192 in a highly conserved Streptococcus pyogenes extracellular cysteine protease (interleukin 1beta convertase) alters proteolytic activity and ablates zymogen processing.

    PubMed

    Musser, J M; Stockbauer, K; Kapur, V; Rudgers, G W

    1996-06-01

    Virtually all strains of the human pathogenic bacterium Streptococcus pyogenes express a highly conserved extracellular cysteine protease. The protein is made as an inactive zymogen of 40,000 Da and undergoes autocatalytic truncation to result in a 28,000-Da active protease. Numerous independent lines of investigation suggest that this enzyme participates in one or more phases of host-parasite interaction, such as inflammation and soft tissue invasion. Replacement of the single cysteine residue (C-192) with serine (C192S mutation) resulted in loss of detectable proteolytic activity against bovine casein, human fibronectin, and the low-molecular-weight synthetic substrate 7-amino-4-trifluoromethyl coumarin. The C192S mutant molecule does not undergo autocatalytic processing of zymogen to mature form. Taken together, these data support the hypothesis that C-192 participates in active-site formation and enzyme catalysis.

  18. Streptococcus pyogenes M49 plasminogen/plasmin binding facilitates keratinocyte invasion via integrin-integrin-linked kinase (ILK) pathways and protects from macrophage killing.

    PubMed

    Siemens, Nikolai; Patenge, Nadja; Otto, Juliane; Fiedler, Tomas; Kreikemeyer, Bernd

    2011-06-17

    The entry into epithelial cells and the prevention of primary immune responses are a prerequisite for a successful colonization and subsequent infection of the human host by Streptococcus pyogenes (group A streptococci, GAS). Here, we demonstrate that interaction of GAS with plasminogen promotes an integrin-mediated internalization of the bacteria into keratinocytes, which is independent from the serine protease activity of potentially generated plasmin. α(1)β(1)- and α(5)β(1)-integrins were identified as the major keratinocyte receptors involved in this process. Inhibition of integrin-linked kinase (ILK) expression by siRNA silencing or blocking of PI3K and Akt with specific inhibitors, reduced the GAS M49-plasminogen/plasmin-mediated invasion of keratinocytes. In addition, blocking of actin polymerization significantly reduced GAS internalization into keratinocytes. Altogether, these results provide a first model of plasminogen-mediated GAS invasion into keratinocytes. Furthermore, we demonstrate that plasminogen binding protects the bacteria against macrophage killing.

  19. The Interaction of Canine Plasminogen with Streptococcus pyogenes Enolase: They Bind to One Another but What Is the Nature of the Structures Involved?

    PubMed Central

    Hancock, Mark A.

    2011-01-01

    For years it has been clear that plasminogen from different sources and enolase from different sources interact strongly. What is less clear is the nature of the structures required for them to interact. This work examines the interaction between canine plasminogen (dPgn) and Streptococcus pyogenes enolase (Str enolase) using analytical ultracentrifugation (AUC), surface plasmon resonance (SPR), fluorescence polarization, dynamic light scattering (DLS), isothermal titration calorimetry (ITC), and simple pull-down reactions. Overall, our data indicate that a non-native structure of the octameric Str enolase (monomers or multimers) is an important determinant of its surface-mediated interaction with host plasminogen. Interestingly, a non-native structure of plasminogen is capable of interacting with native enolase. As far as we can tell, the native structures resist forming stable mixed complexes. PMID:22174817

  20. Phenotypes and genotypes of erythromycin-resistant Streptococcus pyogenes strains in Italy and heterogeneity of inducibly resistant strains.

    PubMed

    Giovanetti, E; Montanari, M P; Mingoia, M; Varaldo, P E

    1999-08-01

    A total of 387 clinical strains of erythromycin-resistant (MIC, >/=1 microg/ml) Streptococcus pyogenes, all isolated in Italian laboratories from 1995 to 1998, were examined. By the erythromycin-clindamycin double-disk test, 203 (52.5%) strains were assigned to the recently described M phenotype, 120 (31.0%) were assigned to the inducible macrolide, lincosamide, and streptogramin B resistance (iMLS) phenotype, and 64 (16.5%) were assigned to the constitutive MLS resistance (cMLS) phenotype. The inducible character of the resistance of the iMLS strains was confirmed by comparing the clindamycin MICs determined under normal testing conditions and those determined after induction by pregrowth in 0.05 microg of erythromycin per ml. The MICs of erythromycin, clarithromycin, azithromycin, josamycin, spiramycin, and the ketolide HMR3004 were then determined and compared. Homogeneous susceptibility patterns were observed for the isolates of the cMLS phenotype (for all but one of the strains, HMR3004 MICs were 0.5 to 8 microg/ml and the MICs of the other drugs were >128 microg/ml) and those of the M phenotype (resistance only to the 14- and 15-membered macrolides was recorded, with MICs of 2 to 32 microg/ml). Conversely, heterogeneous susceptibility patterns were observed in the isolates of the iMLS phenotype, which were subdivided into three distinct subtypes designated iMLS-A, iMLS-B, and iMLS-C. The iMLS-A strains (n = 84) were highly resistant to the 14-, 15-, and 16-membered macrolides and demonstrated reduced susceptibility to low-level resistance to HMR3004. The iMLS-B strains (n = 12) were highly resistant to the 14- and 15-membered macrolides, susceptible to the 16-membered macrolides (but highly resistant to josamycin after induction), and susceptible to HMR3004 (but intermediate or resistant after induction). The iMLS-C strains (n = 24) had lower levels of resistance to the 14- and 15-membered macrolides (with erythromycin MICs increasing two to four times after

  1. Crystal Structure of Streptococcus pyogenes Cas1 and Its Interaction with Csn2 in the Type II CRISPR-Cas System.

    PubMed

    Ka, Donghyun; Lee, Hasup; Jung, Yi-Deun; Kim, Kyunggon; Seok, Chaok; Suh, Nayoung; Bae, Euiyoung

    2016-01-05

    CRISPRs and Cas proteins constitute an RNA-guided microbial immune system against invading nucleic acids. Cas1 is a universal Cas protein found in all three types of CRISPR-Cas systems, and its role is implicated in new spacer acquisition during CRISPR-mediated adaptive immunity. Here, we report the crystal structure of Streptococcus pyogenes Cas1 (SpCas1) in a type II CRISPR-Cas system and characterize its interaction with S. pyogenes Csn2 (SpCsn2). The SpCas1 structure reveals a unique conformational state distinct from type I Cas1 structures, resulting in a more extensive dimerization interface, a more globular overall structure, and a disruption of potential metal-binding sites for catalysis. We demonstrate that SpCas1 directly interacts with SpCsn2, and identify the binding interface and key residues for Cas complex formation. These results provide structural information for a type II Cas1 protein, and lay a foundation for studying multiprotein Cas complexes functioning in type II CRISPR-Cas systems.

  2. Boesenbergia pandurata (Roxb.) Schltr., Eleutherine americana Merr. and Rhodomyrtus tomentosa (Aiton) Hassk. as antibiofilm producing and antiquorum sensing in Streptococcus pyogenes.

    PubMed

    Limsuwan, Surasak; Voravuthikunchai, Supayang Piyawan

    2008-08-01

    Biofilm formation has been demonstrated as a potentially important mechanism contributing to antibiotic treatment failure on Streptococcus pyogenes. It could play a significant role in recurrent and chronic infections. Boesenbergia pandurata (Roxb.) Schltr., Eleutherine americana Merr. and Rhodomyrtus tomentosa (Aiton) Hassk. have been previously reported from our laboratory as effective agents against S. pyogenes. Therefore, in the present study, we observed the effect of these plants on biofilm formation. The bacterial biofilms were quantified by safranin staining and absorbance at 492 nm. The results clearly demonstrated that all subinhibitory concentrations [1/32-1/2 minimal inhibitory concentration (MIC)] of E. americana (7.81-125 microg mL(-1)) and R. tomentosa (0.24-7.81 microg mL(-1)) extracts significantly prevented biofilm formation while 1/2MIC (7.81 microg mL(-1)) of B. pandurata extract produced this effect. The issue of antiquorum sensing of this pathogenic bacterium has been further explored. A correlation between antiquorum-sensing and antibiofilm-producing activities was demonstrated. Strong inhibition on quorum sensing was displayed with the extract of R. tomentosa. Eleutherine americana extract showed partial inhibition, while B. pandurata did not show this activity. By contrast, an assay of microbial adhesion to hydrocarbon revealed no changes in the cell-surface hydrophobicity of the treated organisms. Active organisms with the ability to inhibit quorum sensing and biofilm formation are worth studying as they may provide complimentary medicine for biofilm-associated infections.

  3. Regulation of SpeB in Streptococcus pyogenes by pH and NaCl: a Model for In Vivo Gene Expression†

    PubMed Central

    Loughman, Jennifer A.; Caparon, Michael

    2006-01-01

    For a pathogen such as Streptococcus pyogenes, ecological success is determined by its ability to sense the environment and mount an appropriate adaptive transcriptional response. Thus, determining conditions for analyses of gene expression in vitro that are representative of the in vivo environment is critical for understanding the contributions of transcriptional response pathways to pathogenesis. In this study, we determined that the gene encoding the SpeB cysteine protease is up-regulated over the course of infection in a murine soft-tissue model. Conditions were identified, including growth phase, acidic pH, and an NaCl concentration of <0.1 M, that were required for expression of speB in vitro. Analysis of global expression profiles in response to these conditions in vitro identified a set of coregulated genes whose expression patterns showed a significant correlation with that of speB when examined during infection of murine soft tissues. This analysis revealed that a culture medium that promotes high levels of SpeB expression in vitro produced an expression profile that showed significant correlation to the profile observed in vivo. Taken together, these studies establish culture conditions that mimic in vivo expression patterns; that growth phase, pH, and NaCl may mimic relevant cues sensed by S. pyogenes during infection; and that identification of other environmental cues that alter expression of speB in vitro may provide insight into the signals that direct global patterns of gene expression in vivo. PMID:16385029

  4. Betulin inhibits virulence and biofilm of Streptococcus pyogenes by suppressing ropB core regulon, sagA and dltA.

    PubMed

    Viszwapriya, Dharmaprakash; Subramenium, Ganapathy Ashwinkumar; Prithika, Udayakumar; Balamurugan, Krishnaswamy; Pandian, Shunmugiah Karutha

    2016-10-01

    The present study demonstrates the antivirulence potential of betulin, an abundantly available triterpenoid against Streptococcus pyogenes, a multivirulent and exclusive human pathogen. Crystal violet assay and microscopic examination revealed that betulin (100 μg mL(-1)) exhibits surface-independent antibiofilm activity and mitigates extracellular polymeric substance production. Betulin treatment enhanced the rate of auto-aggregation in liquid medium. Results of real-time PCR and biochemical assays demonstrated that betulin suppresses the expression of ropB core regulon, sagA and dltA, which correspondingly affects SpeB production, hemolysis and cell surface hydrophobicity for the observed impairment in virulence and biofilm formation. dltA downregulation also affected the production of M protein, making betulin-treated cells more susceptible to phagocytosis. The non-toxic nature of betulin and its antivirulence potential against S. pyogenes were manifested in vivo in Caenorhabditis elegans This study reveals the prospective role of betulin as therapeutic agent for the prevention and treatment of streptococcal infections.

  5. Intranasal vaccination with SfbI or M protein-derived peptides conjugated to diphtheria toxoid confers protective immunity against a lethal challenge with Streptococcus pyogenes.

    PubMed

    Schulze, Kai; Olive, Colleen; Ebensen, Thomas; Guzmán, Carlos A

    2006-08-28

    We investigated whether intranasal immunisation with diphtheria toxoid (DT) conjugated polypeptides encompassing T and B cell epitopes of the SfbI protein (FNBR) or a conformational-constrained B cell epitope of the M1 protein (J8) was able to confer protection against lethal mucosal challenge with a heterologous Streptococcus pyogenes strain. To this end, BALB/c mice were immunised with the conjugates. Strong antigen-specific antibody responses were observed in both serum and mucosal secretions. Vaccinated mice were challenged 10 days after the last boost by the intranasal route. Animals receiving FNBR-DT co-administered with either the cholera toxin B subunit (CTB) or the TLR 2/6 agonist MALP-2 were efficiently protected against the virulent S. pyogenes strain (90% and 70% survival, respectively), whereas those immunised with J8-DT plus either CTB or MALP-2 showed intermediate levels of protection (60% and 40%, respectively). The obtained results indicate that in our experimental animal model peptide-based conjugate vaccines represent a valid alternative to protect against streptococcal infection.

  6. Molecular population genetic evidence of horizontal spread of two alleles of the pyrogenic exotoxin C gene (speC) among pathogenic clones of Streptococcus pyogenes.

    PubMed Central

    Kapur, V; Nelson, K; Schlievert, P M; Selander, R K; Musser, J M

    1992-01-01

    It has recently been demonstrated that the bacteriophage-borne gene (speC) encoding pyrogenic exotoxin C is harbored by phylogenetic lineages representing virtually the entire breadth of genomic differentiation present in the species Streptococcus pyogenes (J. M. Musser, A. R. Hauser, M. H. Kim, P. M. Schlievert, K. Nelson, and R. K. Selander, Proc. Natl. Acad. Sci. USA 88:2668-2672, 1991). To determine whether the speC genes occurring in association with divergent chromosomal genotypes (clones) are identical or represent a group of allelic variants, we sequenced speC from 23 S. pyogenes strains representing 15 clones identified by multilocus enzyme electrophoresis. Two alleles of speC are present in natural populations, and each allele occurs in clones that are well differentiated in overall chromosomal character; in one case, isolates of a single clone had different speC alleles. We interpret these patterns of toxin allele-clone distribution as evidence of occasional episodes of speC horizontal dissemination, presumably by bacteriophage-mediated gene transfer and recombination. Images PMID:1500157

  7. Toward New Therapeutics for Skin and Soft Tissue Infections: Propargyl-Linked Antifolates Are Potent Inhibitors of MRSA and Streptococcus pyogenes

    PubMed Central

    Scocchera, Eric W.; Martin, Brooke D.; Swain III, P. Whitney; Alverson, Jeremy B.; Priestley, Nigel D.; Anderson, Amy C.; Wright, Dennis L.

    2012-01-01

    Hospital- and community-acquired, complicated skin and soft tissue infections, often attributed to Staphylococcus aureus and Streptococcus pyogenes, present a significant health burden that is associated with increased health care costs and mortality. As these two species are difficult to discern on diagnosis and are associated with differential profiles of drug resistance, the development of an efficacious antibacterial agent that targets both organisms is a high priority. Herein we describe a structure-based drug development effort that has produced highly potent inhibitors of dihydrofolate reductase from both species. Optimized propargyl-linked antifolates containing a key pyridyl substituent display antibacterial activity against both methicillin-resistant S. aureus and S. pyogenes at MIC values below 0.1 µg/mL and minimal cytotoxicity against mammalian cells. Further evaluation against a panel of clinical isolates shows good efficacy against a range of important phenotypes such as hospital- and community-acquired strains as well as strains resistant to vancomycin. PMID:22347365

  8. Dissemination of the phage-associated novel superantigen gene speL in recent invasive and noninvasive Streptococcus pyogenes M3/T3 isolates in Japan.

    PubMed

    Ikebe, Tadayoshi; Wada, Akihito; Inagaki, Yoshishige; Sugama, Kumiko; Suzuki, Rieko; Tanaka, Daisuke; Tamaru, Aki; Fujinaga, Yoshihiro; Abe, Yoshiaki; Shimizu, Yoshikata; Watanabe, Haruo

    2002-06-01

    In Japan, more than 10% of streptococcal toxic shock-like syndrome (TSLS) cases have been caused by Streptococcus pyogenes M3/T3 isolates since the first reported TSLS case in 1992. Most M3/T3 isolates from TSLS or severe invasive infection cases during 1992 to 2001 and those from noninvasive cases during this period are indistinguishable in pulsed-field gel electropherograms. The longest fragments of these recent isolates were 300 kb in size, whereas those of isolates recovered during or before 1973 were 260 kb in size. These 260- and 300-kb fragments hybridized to each other, suggesting the acquisition of an about 40-kb fragment by the recent isolates. The whole part of the acquired fragment was cloned from the first Japanese TSLS isolate, NIH1, and its nucleotide sequence was determined. The 41,796-bp fragment is temperate phage phiNIH1.1, containing a new superantigen gene speL near its right attachment site. The C-terminal part of the deduced amino acid sequence of speL has 48 and 46% similarity with well-characterized erythrogenic toxin SpeC and the most potent superantigen, SmeZ-2, respectively. None of 10 T3 isolates recovered during or before 1973 has speL, whereas all of 18 M3/T3 isolates recovered during or after 1992 and, surprisingly, Streptococcus equi subsp. equi ATCC 9527 do have this gene. Though plaques could not be obtained from phiNIH1.1, its DNA became detectable from the phage particle fraction upon mitomycin C induction, showing that this phage is not defective. A horizontal transfer of the phage carrying speL may explain the observed change in M3/T3 S. pyogenes isolates in Japan.

  9. Differential Secretomics of Streptococcus pyogenes Reveals a Novel Peroxide Regulator (PerR)-regulated Extracellular Virulence Factor Mitogen Factor3 (MF3)*

    PubMed Central

    Wen, Yao-Tseng; Tsou, Chih-Cheng; Kuo, Hsin-Tzu; Wang, Jie-Siou; Wu, Jiunn-Jong; Liao, Pao-Chi

    2011-01-01

    Streptococcus pyogenes is a human pathogen that causes various diseases. Numerous virulence factors secreted by S. pyogenes are involved in pathogenesis. The peroxide regulator (PerR) is associated with the peroxide resistance response and pathogenesis, but little is known about the regulation of the secretome involved in virulence. To investigate how PerR regulates the expression of the S. pyogenes secretome involved in virulence, a perR deficient mutant was used for comparative secretomic analysis with a wild-type strain. The conditioned medium containing secreted proteins of a wild-type strain and a perR deficient mutant at the stationary phase were collected for two-dimensional gel electrophoresis analysis, where protease inhibitors were applied to avoid the degradation of extracellular proteins. Differentially expressed protein spots were identified by liquid chromatography electrospray ionization tandem MS. More than 330 protein spots were detected on each gel. We identified 25 unique up-regulated proteins and 13 unique down-regulated proteins that were directly or indirectly controlled by the PerR regulator. Among these identified proteins, mitogen factor 3 (MF3), was selected to verify virulence and the expression of gene products. The data showed that MF3 protein levels in conditioned medium, as measured by immunoblot analysis, correlated well with protein levels determined by two-dimensional gel electrophoresis analysis. We also demonstrated that PerR bound to the promoter region of the mf3 gene. The result of an infection model showed that virulence was attenuated in the mf3 deficient mutant. Additional growth data of the wild-type strain and the mf3 deficient mutant suggested that MF3 played a role in digestion of exogenous DNA for promoting growth. To summarize, we conclude that PerR can positively regulate the expression of the secreted protein MF3 that contributes to the virulence in S. pyogenes. The analysis of the PerR-regulated secretome provided

  10. SpyB, a small heme-binding protein, affects the composition of the cell wall in Streptococcus pyogenes

    SciTech Connect

    Edgar, Rebecca J.; Chen, Jing; Kant, Sashi; Rechkina, Elena; Rush, Jeffrey S.; Forsberg, Lennart S.; Jaehrig, Bernhard; Azadi, Parastoo; Tchesnokova, Veronika; Sokurenko, Evgeni V.; Zhu, Haining; Korotkov, Konstantin V.; Pancholi, Vijay; Korotkova, Natalia

    2016-10-13

    Streptococcus pyogenes (Group A Streptococcus or GAS) is a hemolytic human pathogen associated with a wide variety of infections ranging from minor skin and throat infections to life-threatening invasive diseases. The cell wall of GAS consists of peptidoglycan sacculus decorated with a carbohydrate comprising a polyrhamnose backbone with immunodominant N-acetylglucosamine side-chains. All GAS genomes contain the spyBA operon, which encodes a 35-amino-acid membrane protein SpyB, and a membrane-bound C3-like ADP-ribosyltransferase SpyA. In this study, we addressed the function of SpyB in GAS. Phenotypic analysis of a spyB deletion mutant revealed increased bacterial aggregation, and reduced sensitivity to β-lactams of the cephalosporin class and peptidoglycan hydrolase PlyC. Glycosyl composition analysis of cell wall isolated from the spyB mutant suggested an altered carbohydrate structure compared with the wild-type strain. Furthermore, we found that SpyB associates with heme and protoporphyrin IX. Heme binding induces SpyB dimerization, which involves disulfide bond formation between the subunits. Furthermore, our data suggest the possibility that SpyB activity is regulated by heme.

  11. Long-term antibody memory induced by synthetic peptide vaccination is protective against Streptococcus pyogenes infection and is independent of memory T cell help.

    PubMed

    Pandey, Manisha; Wykes, Michelle N; Hartas, Jon; Good, Michael F; Batzloff, Michael R

    2013-03-15

    Streptococcus pyogenes (group A Streptococcus [GAS]) is a leading human pathogen associated with a diverse array of mucosal and systemic infections. Vaccination with J8, a conserved region synthetic peptide derived from the M-protein of GAS and containing only 12 aa from GAS, when conjugated to diphtheria toxoid, has been shown to protect mice against a lethal GAS challenge. Protection has been previously shown to be Ab-mediated. J8 does not contain a dominant GAS-specific T cell epitope. The current study examined long-term Ab memory and dissected the role of B and T cells. Our results demonstrated that vaccination generates specific memory B cells (MBC) and long-lasting Ab responses. The MBC response can be activated following boost with Ag or limiting numbers of whole bacteria. We further show that these memory responses protect against systemic infection with GAS. T cell help is required for activation of MBC but can be provided by naive T cells responding directly to GAS at the time of infection. Thus, individuals whose T cells do not recognize the short synthetic peptide in the vaccine will be able to generate a protective and rapid memory Ab response at the time of infection. These studies significantly strengthen previous findings, which showed that protection by the J8-diphtheria toxoid vaccine is Ab-mediated and suggest that in vaccine design for other organisms the source of T cell help for Ab responses need not be limited to sequences from the organism itself.

  12. The PerR-regulated P1B-4 type ATPase (PmtA) acts as a ferrous iron efflux pump in Streptococcus pyogenes.

    PubMed

    Turner, Andrew G; Ong, Cheryl-Lynn Y; Djoko, Karrera Y; West, Nicholas P; Davies, Mark R; McEwan, Alastair G; Walker, Mark J

    2017-04-03

    Streptococcus pyogenes (group A Streptococcus; GAS) is an obligate human pathogen responsible for a broad spectrum of human disease. GAS has a requirement for metal homeostasis within the human host and as such, tightly modulates metal uptake and efflux during infection. Metal acquisition systems are required to combat metal sequestration by the host, while metal efflux systems are essential to protect against metal overload poisoning. Here, we investigated the function of PmtA (PerR-regulated metal transporter A), a P1B-4 type ATPase efflux pump, in the invasive GAS M1T1 strain 5448. We reveal that PmtA functions as a ferrous iron [Fe(II)] efflux system. In the presence of high Fe(II) concentrations, the 5448ΔpmtA deletion mutant exhibited diminished growth and accumulated 5-fold higher intracellular Fe(II) compared to the wild-type and complemented mutant. The 5448ΔpmtA deletion mutant also showed enhanced susceptibility to killing by the Fe-dependent antibiotic, streptonigrin, as well as increased sensitivity to hydrogen peroxide and superoxide. We suggest that the PerR-mediated control of Fe(II) efflux by PmtA is important for bacterial defense against oxidative stress. PmtA represents an exemplar for a Fe(II) efflux system in a host-adapted Gram-positive bacterial pathogen.

  13. Interaction between complement regulators and Streptococcus pyogenes: binding of C4b-binding protein and factor H/factor H-like protein 1 to M18 strains involves two different cell surface molecules.

    PubMed

    Pérez-Caballero, David; García-Laorden, Isabel; Cortés, Guadalupe; Wessels, Michael R; de Córdoba, Santiago Rodríguez; Albertí, Sebastián

    2004-12-01

    Streptococcus pyogenes, or group A Streptococcus, is one of the most frequent causes of pharyngitis and skin infections in humans. Many virulence mechanisms have been suggested to be involved in the infectious process. Among them is the binding to the bacterial cell surface of the complement regulatory proteins factor H, factor H-like protein 1 (FHL-1), and C4b-binding protein. Previous studies indicate that binding of these three regulators to the streptococcal cell involves the M protein encoded by the emm gene. M-type 18 strains are prevalent among clinical isolates and have been shown to interact with all three complement regulators simultaneously. Using isogenic strains lacking expression of the Emm18 or the Enn18 proteins, we demonstrate in this study that, in contradistinction to previously described S. pyogenes strains, M18 strains bind the complement regulators factor H, FHL-1, and C4b-binding protein through two distinct cell surface proteins. Factor H and FHL-1 bind to the Emm18 protein, while C4BP binds to the Enn18 protein. We propose that expression of two distinct surface structures that bind complement regulatory proteins represents a unique adaptation of M18 strains that enhances their resistance to opsonization by human plasma and increases survival of this particular S. pyogenes strain in the human host. These new findings illustrate that S. pyogenes has evolved diverse mechanisms for recruitment of complement regulatory proteins to the bacterial surface to evade immune clearance in the human host.

  14. SpxA1 and SpxA2 Act Coordinately To Fine-Tune Stress Responses and Virulence in Streptococcus pyogenes

    PubMed Central

    Port, Gary C.; Cusumano, Zachary T.; Tumminello, Paul R.

    2017-01-01

    ABSTRACT SpxA is a unique transcriptional regulator highly conserved among members of the phylum Firmicutes that binds RNA polymerase and can act as an antiactivator. Why some Firmicutes members have two highly similar SpxA paralogs is not understood. Here, we show that the SpxA paralogs of the pathogen Streptococcus pyogenes, SpxA1 and SpxA2, act coordinately to regulate virulence by fine-tuning toxin expression and stress resistance. Construction and analysis of mutants revealed that SpxA1− mutants were defective for growth under aerobic conditions, while SpxA2− mutants had severely attenuated responses to multiple stresses, including thermal and oxidative stresses. SpxA1− mutants had enhanced resistance to the cationic antimicrobial molecule polymyxin B, while SpxA2− mutants were more sensitive. In a murine model of soft tissue infection, a SpxA1− mutant was highly attenuated. In contrast, the highly stress-sensitive SpxA2− mutant was hypervirulent, exhibiting more extensive tissue damage and a greater bacterial burden than the wild-type strain. SpxA1− attenuation was associated with reduced expression of several toxins, including the SpeB cysteine protease. In contrast, SpxA2− hypervirulence correlated with toxin overexpression and could be suppressed to wild-type levels by deletion of speB. These data show that SpxA1 and SpxA2 have opposing roles in virulence and stress resistance, suggesting that they act coordinately to fine-tune toxin expression in response to stress. SpxA2− hypervirulence also shows that stress resistance is not always essential for S. pyogenes pathogenesis in soft tissue. PMID:28351920

  15. tRNA modification by GidA/MnmE is necessary for Streptococcus pyogenes virulence: a new strategy to make live attenuated strains.

    PubMed

    Cho, Kyu Hong; Caparon, Michael G

    2008-07-01

    Studies directed at vaccine development and mucosal immunity against Streptococcus pyogenes would benefit from the availability of live attenuated strains. Our approach for production of candidate live attenuated strains was to identify mutations that did not alter growth in vitro and did not alter the overall complement of virulence factors produced but did result in reduced levels of expression of multiple secreted virulence factors. A global reduction but not elimination of expression would likely lead to attenuation while maximizing the number of antigenic targets available for stimulation of immunity. Adaptation of Tn5-based transposome mutagenesis to S. pyogenes with initial screening for reduced expression of the SpeB protease resulted in identification of mutations in gidA, which encodes an enzyme involved in tRNA modification. Reduced SpeB expression was due to delayed onset of speB transcription resulting from reduced translation efficiency of the message for RopB, a transcriptional activator. Overall, GidA(-) mutants had a nearly normal global transcription profile but expressed significantly reduced levels of multiple virulence factors due to impaired translation efficiencies. A translation defect was supported by the observation that mutants lacking MnmE, which functions in the same tRNA modification pathway as GidA, phenocopied GidA deficiency. The mutants stimulated a cytokine response in cultured macrophages identical to that in the wild type, with the exception of reduced levels of tumor necrosis factor alpha and interleukin-23. Significantly, GidA(-) mutants were highly attenuated in the murine ulcer model of soft tissue infection. These characteristics suggest that GidA pathway tRNA modification mutants are attractive candidates for further evaluation as live attenuated strains.

  16. What causes decreased erythromycin resistance in Streptococcus pyogenes? Dynamics of four clones in a southern European region from 2005 to 2012.

    PubMed

    Montes, Milagrosa; Tamayo, Esther; Mojica, Catalina; García-Arenzana, José M; Esnal, Olatz; Pérez-Trallero, Emilio

    2014-06-01

    To survey antibiotic resistance among Streptococcus pyogenes isolates collected from 2005 to 2012, to characterize those showing erythromycin resistance and to analyse the association of certain emm types with erythromycin resistance or susceptibility. Resistance determinants or mutations conferring erythromycin, clindamycin, tetracycline and fluoroquinolone resistance were analysed. All erythromycin-resistant isolates and a sample of erythromycin-susceptible isolates were emm typed. Multilocus sequence typing was performed for representative emm types. Antimicrobial susceptibility was studied for 12 346 S. pyogenes isolates. Erythromycin, clindamycin and tetracycline resistance showed a decreasing trend. In 2012, 2.8% of isolates were erythromycin resistant versus 7.5% in 2005 and 11.7% in 2006. Although 21 clones were involved, 4 clones accounted for almost 90% of erythromycin-resistant isolates. The emm12/ST36 clone, carrying the mef(A) gene, was the predominant (41.1%) erythromycin-resistant clone, with an incidence peak in 2008, followed by a gradual decline. The M phenotype predominated each year except for 2005, when two of the main erythromycin-resistant clones (emm11/ST403 and emm28/ST52) harboured an erm(B) gene. Erythromycin resistance was significantly higher in adults than in children. Skin isolates showed the highest erythromycin resistance rate; among these, perianal isolates frequently belonged to the emm28/ST52 clone. The emm type was not a predictor of erythromycin resistance; however, most emm11 and emm12 were erythromycin-resistant isolates. Macrolide consumption was similar throughout the study period. Only two isolates with a high level of levofloxacin resistance were detected. Resistance was mainly related to the circulation of emm12/ST36, emm11/ST403, emm28/ST52 and emm4/ST39 clones, all of which declined throughout the study period. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial

  17. The Membrane Bound LRR Lipoprotein Slr, and the Cell Wall-Anchored M1 Protein from Streptococcus pyogenes Both Interact with Type I Collagen

    PubMed Central

    Bober, Marta; Mörgelin, Matthias; Olin, Anders I.; von Pawel-Rammingen, Ulrich; Collin, Mattias

    2011-01-01

    Streptococcus pyogenes is an important human pathogen and surface structures allow it to adhere to, colonize and invade the human host. Proteins containing leucine rich repeats (LRR) have been indentified in mammals, viruses, archaea and several bacterial species. The LRRs are often involved in protein-protein interaction, are typically 20–30 amino acids long and the defining feature of the LRR motif is an 11-residue sequence LxxLxLxxNxL (x being any amino acid). The streptococcal leucine rich (Slr) protein is a hypothetical lipoprotein that has been shown to be involved in virulence, but at present no ligands for Slr have been identified. We could establish that Slr is a membrane attached horseshoe shaped lipoprotein by homology modeling, signal peptidase II inhibition, electron microscopy (of bacteria and purified protein) and immunoblotting. Based on our previous knowledge of LRR proteins we hypothesized that Slr could mediate binding to collagen. We could show by surface plasmon resonance that recombinant Slr and purified M1 protein bind with high affinity to collagen I. Isogenic slr mutant strain (MB1) and emm1 mutant strain (MC25) had reduced binding to collagen type I as shown by slot blot and surface plasmon resonance. Electron microscopy using gold labeled Slr showed multiple binding sites to collagen I, both to the monomeric and the fibrillar structure, and most binding occurred in the overlap region of the collagen I fibril. In conclusion, we show that Slr is an abundant membrane bound lipoprotein that is co-expressed on the surface with M1, and that both these proteins are involved in recruiting collagen type I to the bacterial surface. This underlines the importance of S. pyogenes interaction with extracellular matrix molecules, especially since both Slr and M1 have been shown to be virulence factors. PMID:21655249

  18. Phenotypes and genotypes of erythromycin-resistant Streptococcus pyogenes strains isolated from invasive and non-invasive infections from Mexico and the USA during 1999-2010.

    PubMed

    Villaseñor-Sierra, Alberto; Katahira, Eva; Jaramillo-Valdivia, Abril N; Barajas-García, María de los Angeles; Bryant, Amy; Morfín-Otero, Rayo; Márquez-Díaz, Francisco; Tinoco, Juan Carlos; Sánchez-Corona, José; Stevens, Dennis L

    2012-03-01

    To compare the prevalence, phenotypes, and genes responsible for erythromycin resistance among Streptococcus pyogenes isolates from Mexico and the USA. Eighty-nine invasive and 378 non-invasive isolates from Mexico, plus 148 invasive, 21 non-invasive, and five unclassified isolates from the USA were studied. Susceptibilities to penicillin, erythromycin, clindamycin, ceftriaxone, and vancomycin were evaluated according to Clinical and Laboratory Standards Institute (CLSI) standards. Phenotypes of erythromycin resistance were identified by triple disk test, and screening for mefA, ermTR, and ermB genes was carried out by PCR. All isolates were susceptible to penicillin, ceftriaxone, and vancomycin. Erythromycin resistance was found in 4.9% of Mexican strains and 5.2% of USA strains. Phenotypes in Mexican strains were 95% M and 5% cMLS; in strains from the USA, phenotypes were 33.3% iMLS, 33.3% iMLS-D, and 33.3% M. Erythromycin resistance genes in strains from Mexico were mefA (95%) and ermB (5%); USA strains harbored ermTR (56%), mefA (33%), and none (11%). In Mexico, all erythromycin-resistant strains were non-invasive, whereas 89% of strains from the USA were invasive. Erythromycin resistance continues to exist at low levels in both Mexico and the USA, although the genetic mechanisms responsible differ between the two nations. These genetic differences may be related to the invasive character of the S. pyogenes isolated. Copyright © 2011 International Society for Infectious Diseases. All rights reserved.

  19. Phenotypes and genotypes of erythromycin-resistant Streptococcus pyogenes strains isolated from invasive and non-invasive infections from Mexico and the USA during 1999–2010

    PubMed Central

    Villaseñor-Sierra, Alberto; Katahira, Eva; Jaramillo-Valdivia, Abril N.; de los Angeles Barajas-García, María; Bryant, Amy; Morfín-Otero, Rayo; Márquez-Díaz, Francisco; Tinoco, Juan Carlos; Sánchez-Corona, José; Stevens, Dennis L.

    2012-01-01

    Summary Objective To compare the prevalence, phenotypes, and genes responsible for erythromycin resistance among Streptococcus pyogenes isolates from Mexico and the USA. Methods Eighty-nine invasive and 378 non-invasive isolates from Mexico, plus 148 invasive, 21 non-invasive, and five unclassified isolates from the USA were studied. Susceptibilities to penicillin, erythromycin, clindamycin, ceftriaxone, and vancomycin were evaluated according to Clinical and Laboratory Standards Institute (CLSI) standards. Phenotypes of erythromycin resistance were identified by triple disk test, and screening for mefA, ermTR, and ermB genes was carried out by PCR. Results All isolates were susceptible to penicillin, ceftriaxone, and vancomycin. Erythromycin resistance was found in 4.9% of Mexican strains and 5.2% of USA strains. Phenotypes in Mexican strains were 95% M and 5% cMLS; in strains from the USA, phenotypes were 33.3% iMLS, 33.3% iMLS-D, and 33.3% M. Erythromycin resistance genes in strains from Mexico were mefA (95%) and ermB (5%); USA strains harbored ermTR (56%), mefA (33%), and none (11%). In Mexico, all erythromycin-resistant strains were non-invasive, whereas 89% of strains from the USA were invasive. Conclusions Erythromycin resistance continues to exist at low levels in both Mexico and the USA, although the genetic mechanisms responsible differ between the two nations. These genetic differences may be related to the invasive character of the S. pyogenes isolated. PMID:22217469

  20. CovRS-Regulated Transcriptome Analysis of a Hypervirulent M23 Strain of Group A Streptococcus pyogenes Provides New Insights into Virulence Determinants

    PubMed Central

    Bao, Yun-Juan; Liang, Zhong; Mayfield, Jeffrey A.; Lee, Shaun W.; Ploplis, Victoria A.

    2015-01-01

    ABSTRACT The two-component control of virulence (Cov) regulator (R)-sensor (S) (CovRS) regulates the virulence of Streptococcus pyogenes (group A Streptococcus [GAS]). Inactivation of CovS during infection switches the pathogenicity of GAS to a more invasive form by regulating transcription of diverse virulence genes via CovR. However, the manner in which CovRS controls virulence through expression of extended gene families has not been fully determined. In the current study, the CovS-regulated gene expression profiles of a hypervirulent emm23 GAS strain (M23ND/CovS negative [M23ND/CovS−]) and a noninvasive isogenic strain (M23ND/CovS+), under different growth conditions, were investigated. RNA sequencing identified altered expression of ∼349 genes (18% of the chromosome). The data demonstrated that M23ND/CovS− achieved hypervirulence by allowing enhanced expression of genes responsible for antiphagocytosis (e.g., hasABC), by abrogating expression of toxin genes (e.g., speB), and by compromising gene products with dispensable functions (e.g., sfb1). Among these genes, several (e.g., parE and parC) were not previously reported to be regulated by CovRS. Furthermore, the study revealed that CovS also modulated the expression of a broad spectrum of metabolic genes that maximized nutrient utilization and energy metabolism during growth and dissemination, where the bacteria encounter large variations in available nutrients, thus restructuring metabolism of GAS for adaption to diverse growth environments. From constructing a genome-scale metabolic model, we identified 16 nonredundant metabolic gene modules that constitute unique nutrient sources. These genes were proposed to be essential for pathogen growth and are likely associated with GAS virulence. The genome-wide prediction of genes associated with virulence identifies new candidate genes that potentially contribute to GAS virulence. IMPORTANCE The CovRS system modulates transcription of ∼18% of the genes in

  1. Streptococcus pyogenes-induced cutaneous lymphocyte antigen-positive T cell-dependent epidermal cell activation triggers TH17 responses in patients with guttate psoriasis.

    PubMed

    Ruiz-Romeu, Ester; Ferran, Marta; Sagristà, Marc; Gómez, Julià; Giménez-Arnau, Ana; Herszenyi, Krisztina; Hóllo, Péter; Celada, Antonio; Pujol, Ramon; Santamaria-Babí, Luis F

    2016-08-01

    Guttate psoriasis (GP) is characterized by acute onset of small, rounded psoriatic lesions. Although this particular phenotype of psoriasis is usually associated with streptococcal throat infections and mainly occurs in HLA-Cw6(+) patients, the specific immunologic response to this innate stimulus that causes these skin lesions is poorly understood. This study aims to elucidate how key cellular elements of patients with GP respond to Streptococcus pyogenes and whether this initial immune response is favored by the genetic and environmental background of these patients. Circulating memory T cells and autologous epidermal cells from samples from either patients with GP (n = 14) or healthy control subjects (n = 6) were cocultured ex vivo in the presence of an S pyogenes extract. Levels of the psoriasis-associated cytokines IL-17A, IL-17F, IFN-γ, TNF-α, IL-6, and IL-8 were determined. The expression of several genes with increased (DEFB4, S100A7, LCN2, IL36G, IL8, CXCL9, CXCL10, and CXCL11) or decreased (FLG and LOR) transcripts in psoriatic lesions was examined in keratinocytes treated with coculture supernatants. When skin-homing effector memory cutaneous lymphocyte antigen-positive T cells were used in cocultures, a TH17-dominant response was observed, as reflected by the higher amounts of IL-17A and IL-17F than IFN-γ. Moreover, a higher TH17 response was observed in cells isolated from patients with flares associated with a streptococcal tonsillitis and with the HLA-Cw6 allele (cohort 1). In addition, in normal keratinocytes the supernatants from these cocultures induced an increase in IL-17-associated genes, such as DEFB4, S100A7, LCN2, IL36G, and IL8 but a decrease in FLG and LOR, thereby confirming the role of activated TH17 cells. This study reveals a dominant TH17 response of cutaneous lymphocyte antigen-positive T cells activated by epidermal cells and S pyogenes in patients with GP. Copyright © 2016 American Academy of Allergy, Asthma

  2. Pyogenic granuloma

    MedlinePlus

    ... page: //medlineplus.gov/ency/article/001464.htm Pyogenic granuloma To use the sharing features on this page, please enable JavaScript. Pyogenic granulomas are small, raised, and red bumps on the ...

  3. Evaluation of the antimicrobial efficacy of green tea extract (egcg) against streptococcus pyogenes in vitro - biomed 2011.

    PubMed

    Hull Vance, Stacy; Tucci, Michelle; Benghuzzi, Hamed

    2011-01-01

    S. pyogenes (GAS) is the agent that is involved in acute infections that can lead to devastating life-threatening invasive infections. S. pyogenes cell-surface components such as lipoteichoic acid, hyaluronic acid capsule, M proteins, and laminin and collagen binding proteins are responsible for adhesion to human cells. Natural compounds have been associated with decreased adherence of bacteria to cells. The most widely studies of these compounds has been cranberry juice which is thought to disrupt cellular adherence by altering the local cellular. The purpose of this study was to determine if pre-treating or post-treating kidney epithelial cells (RMKEC) with Epigallocatechin Gallate (EGCG) alters GAS’s ability to adhere to the host cells. Our data indicates that pretreating or post treating the cells with EGCG (10µM, 30µM, or 50µM) inhibited the attachment of the bacteria to the cells in a dose dependent manner. The results also indicate that the remaining GAS bacteria attached to the cellular surface after a 24 hour period was unable to penetrate the epithelial cells. Additional studies also showed EGCG has antimicrobial properties at various concentrations. Our results indicate that EGCG can effectively reduce GAS bacteria cellular attachment and induce GAS cell death and can be used effectively as an adjunct to conventional antibiotic treatments.

  4. Case report: Co-infection of Rickettsia rickettsii and Streptococcus pyogenes: is fatal Rocky Mountain spotted fever underdiagnosed?

    PubMed

    Raczniak, Gregory A; Kato, Cecilia; Chung, Ida H; Austin, Amy; McQuiston, Jennifer H; Weis, Erica; Levy, Craig; Carvalho, Maria da Gloria S; Mitchell, Audrey; Bjork, Adam; Regan, Joanna J

    2014-12-01

    Rocky Mountain spotted fever, a tick-borne disease caused by Rickettsia rickettsii, is challenging to diagnose and rapidly fatal if not treated. We describe a decedent who was co-infected with group A β-hemolytic streptococcus and R. rickettsii. Fatal cases of Rocky Mountain spotted fever may be underreported because they present as difficult to diagnose co-infections.

  5. Regulation of streptokinase expression by CovR/S in Streptococcus pyogenes: CovR acts through a single high-affinity binding site.

    PubMed

    Churchward, Gordon; Bates, Christopher; Gusa, Asiya A; Stringer, Virginia; Scott, June R

    2009-02-01

    The important human pathogen Streptococcus pyogenes (the group A streptococcus or GAS) produces many virulence factors that are regulated by the two-component signal transduction system CovRS (CsrRS). Dissemination of GAS infection originating at the skin has been shown to require production of streptokinase, whose transcription is repressed by CovR. In this work we have studied the interaction of CovR and phosphorylated CovR (CovR-P) with the promoter for streptokinase, Pska. We found that, in contrast to the other CovR-repressed promoters, Pska regulation by CovR occurs through binding at a single ATTARA consensus binding sequence (CB) that overlaps the -10 region of the promoter. Binding of CovR to other nearby consensus sequences occurs upon phosphorylation of the protein, but these other CBs do not contribute to the regulation of Pska by CovR. Thus, binding at a specific site does not necessarily indicate that the site is involved in regulation by CovR. In addition, at Pska, CovR binding to the different sites does not appear to involve cooperative interactions, which simplifies the analysis of CovR binding and gives us insight into the modes of interaction that occur between CovR and its specific DNA-binding sites. Finally, the observation that regulation of transcription from Pska occurs at a very low concentration of phosphorylated CovR may have important implications for the regulation of virulence gene expression during GAS infection.

  6. A role for trigger factor and an rgg-like regulator in the transcription, secretion and processing of the cysteine proteinase of Streptococcus pyogenes.

    PubMed Central

    Lyon, W R; Gibson, C M; Caparon, M G

    1998-01-01

    The ability of numerous microorganisms to cause disease relies upon the highly regulated expression of secreted proteinases. In this study, mutagenesis with a novel derivative of Tn4001 was used to identify genes required for the expression of the secreted cysteine proteinase (SCP) of the pathogenic Gram-positive bacterium Streptococcus pyogenes. Designated as Rop loci (regulation of proteinase), ropB is a rgg-like transcriptional activator required for transcription of the gene which encodes the proteinase. In contrast, ropA contributes post-transcriptionally to the secretion and processing of SCP and encodes a homologue of Trigger Factor, a peptidyl-prolyl isomerase and putative chaparone which is highly conserved in most bacterial species, but of unknown function. Analysis of additional ropA mutants demonstrated that RopA acts both to assist in targeting SCP to the secretory pathway and to promote the ability of the proprotein to establish an active conformation upon secretion. This latter function was dependent upon the peptidyl-prolyl isomerase domain of RopA and mutants that lacked this domain exhibited a bipartite deficiency manifested as a kinetic defect in autologous processing of the proprotein to the mature proteinase, and as a catalytic defect in the mature proteinase. These results provide insight into the function of Trigger Factor, the regulation of proteinase activity and the mechanism of secretion in Gram-positive bacteria. PMID:9799235

  7. Antibacterial resistance in Streptococcus pyogenes (GAS) from healthy carriers and tonsillitis patients and association with antibacterial sale in the Faroe Islands.

    PubMed

    Magnussen, Marita D; Gaini, Shahin; Gislason, Hannes; Kristinsson, Karl G

    2016-04-01

    The aim of this study was to investigate the antibacterial resistance of Streptococcus pyogenes (GAS), and correlate the findings with the sales of erythromycin and tetracycline. General practitioners in the Faroe Islands were recruited to send oropharyngeal swabs. From an ongoing pneumococcal study, nasopharyngeal swabs were sampled from healthy children 0-7 years of age. Erythromycin susceptibility data from Iceland were obtained from the reference laboratory at the Landspitali University Hospital. Susceptibility testing in the Faroe Islands and Iceland was performed according to CLSI methods and criteria. The resistance rate to erythromycin and tetracycline found in patients in the Faroe Islands in 2009/2010 was 6% and 30% respectively. Tetracycline resistance in patients declined significantly from 2009 to 2010 (37-10%, p-value = 0.006 < 0.05) and differed significantly between age groups (p-value = 0.03 < 0.05). In Iceland, there was a peak in erythromycin resistance in 2008 (44%) and a substantial decrease in 2009 (5%). Although the prevalence of erythromycin and tetracycline resistance in the Faroe Islands and Iceland may be associated with antimicrobial use, sudden changes can occur with the introduction of new resistant clones.

  8. Ultrahigh and High Resolution Structures and Mutational Analysis of Monomeric Streptococcus pyogenes SpeB Reveal a Functional Role for the Glycine-rich C-terminal Loop

    SciTech Connect

    González-Páez, Gonzalo E.; Wolan, Dennis W.

    2012-09-05

    Cysteine protease SpeB is secreted from Streptococcus pyogenes and has been studied as a potential virulence factor since its identification almost 70 years ago. Here, we report the crystal structures of apo mature SpeB to 1.06 {angstrom} resolution as well as complexes with the general cysteine protease inhibitor trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane and a novel substrate mimetic peptide inhibitor. These structures uncover conformational changes associated with maturation of SpeB from the inactive zymogen to its active form and identify the residues required for substrate binding. With the use of a newly developed fluorogenic tripeptide substrate to measure SpeB activity, we determined IC{sub 50} values for trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane and our new peptide inhibitor and the effects of mutations within the C-terminal active site loop. The structures and mutational analysis suggest that the conformational movements of the glycine-rich C-terminal loop are important for the recognition and recruitment of biological substrates and release of hydrolyzed products.

  9. SpyA, a C3-Like ADP-Ribosyltransferase, Contributes to Virulence in a Mouse Subcutaneous Model of Streptococcus pyogenes Infection ▿ † ‡

    PubMed Central

    Hoff, Jessica S.; DeWald, Mark; Moseley, Steve L.; Collins, Carleen M.; Voyich, Jovanka M.

    2011-01-01

    Streptococcus pyogenes is an important human pathogen with an expansive repertoire of verified and putative virulence factors. Here we demonstrate that a mutant deficient in the production of the streptococcal ADP-ribosyltransferase SpyA generates lesions of reduced size in a subcutaneous mouse infection model. At early stages of infection, when the difference in lesion size is first established, inflamed tissue isolated from lesions of mice infected with spyA mutant bacteria has higher levels of mRNA encoding the chemokines CXCL1 and CCL2 than does tissue isolated from mice infected with wild-type bacteria. In addition, at these early times, the mRNA levels for the gene encoding the intermediate filament vimentin are higher in the mutant-infected tissue. As wound resolution progresses, mRNA levels of the gene encoding matrix metallopeptidase 2 are lower in mutant-infected tissue. Furthermore, we demonstrate that the spyA mutant is internalized more efficiently than wild-type bacteria by HeLa cells. We conclude that SpyA contributes to streptococcal pathogenesis in the mouse subcutaneous infection model. Our observations suggest that the presence of SpyA delays wound healing in this model. PMID:21422178

  10. Purification, crystallization and preliminary X-ray analysis of native and selenomethionine class I tagatose-1,6-bisphosphate aldolase from Streptococcus pyogenes.

    PubMed

    Liotard, Brigitte; Sygusch, Jurgen

    2004-03-01

    Tagatose-1,6-bisphosphate aldolase (EC 4.1.2.40) is situated at the branching of the tagatose-6-phosphate and Embden-Meyerhof-Parnas (glycolysis) metabolic pathways, where it catalyzes the reversible cleavage of tagatose-1,6-bisphosphate to dihydroxyacetone phosphate and glyceraldehyde 3-phosphate. The recombinant protein from Streptococcus pyogenes was overexpressed in Escherichia coli in its native and selenomethionine-derivative forms and purified using ion-exchange and hydrophobic interaction chromatography. Orthorhombic crystals suitable for structural analysis were obtained by the hanging-drop vapour-diffusion method for both isoforms. The crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 63.7, b = 108.1, c = 238.7 A for the native form and a = 64.1, b = 108.3, c = 239.8 A for the selenomethionine derivative. The asymmetric unit contains four protomers, corresponding to a crystal volume per protein weight (V(M)) of 2.8 A(3) Da(-1) and a solvent content of 56% by volume.

  11. IgG-degrading enzyme of Streptococcus pyogenes (IdeS) prevents disease progression and facilitates improvement in a rabbit model of Guillain-Barré syndrome.

    PubMed

    Wang, Yuzhong; Shi, Qiguang; Lv, Hui; Hu, Ming; Wang, Weifang; Wang, Quanquan; Qiao, Baojun; Zhang, Guorong; Lv, Zhanyun; Kjellman, Christian; Järnum, Sofia; Winstedt, Lena; Zhang, Yong; Wen, Jiao; Hao, Yanlei; Yuki, Nobuhiro

    2017-05-01

    Autoantibodies binding to peripheral nerves followed by complement deposition and membrane attack complex formation results in nerve damage in Guillain-Barré syndrome (GBS). Strategies to remove the pathogenic autoantibodies or block the complement deposition benefit most patients with GBS. Immunoglobulin G-degrading enzyme of Streptococcus pyogenes (IdeS) is a cysteine protease which cleaves IgG antibodies into F(ab')2 and Fc fragments. In this study, using a rabbit model of axonal GBS, acute motor axonal neuropathy (AMAN), we demonstrated that IdeS treatment significantly reduced the disruption of Nav channels as well as activated C3 deposition at the anterior spinal root nodes of Ranvier in AMAN rabbits. IdeS significantly promoted the clinical recovery of AMAN rabbits and there were significant lower frequencies of axonal degeneration in anterior spinal roots of AMAN rabbits with IdeS treatment compared to the saline controls. Our data support that IdeS treatment is a promising therapeutic strategy for GBS. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. A Streptococcus pyogenes derived collagen-like protein as a non-cytotoxic and non-immunogenic cross-linkable biomaterial.

    PubMed

    Peng, Yong Y; Yoshizumi, Ayumi; Danon, Stephen J; Glattauer, Veronica; Prokopenko, Olga; Mirochnitchenko, Oleg; Yu, Zhuoxin; Inouye, Masayori; Werkmeister, Jerome A; Brodsky, Barbara; Ramshaw, John A M

    2010-04-01

    A range of bacteria have been shown to contain collagen-like sequences that form triple-helical structures. Some of these proteins have been shown to form triple-helical motifs that are stable around body temperature without the inclusion of hydroxyproline or other secondary modifications to the protein sequence. This makes these collagen-like proteins particularly suitable for recombinant production as only a single gene product and no additional enzyme needs to be expressed. In the present study, we have examined the cytotoxicity and immunogenicity of the collagen-like domain from Streptococcus pyogenes Scl2 protein. These data show that the purified, recombinant collagen-like protein is not cytotoxic to fibroblasts and does not elicit an immune response in SJL/J and Arc mice. The freeze dried protein can be stabilised by glutaraldehyde cross-linking giving a material that is stable at >37 degrees C and which supports cell attachment while not causing loss of viability. These data suggest that bacterial collagen-like proteins, which can be modified to include specific functional domains, could be a useful material for medical applications and as a scaffold for tissue engineering.

  13. SmaI macrorestriction analysis of Italian isolates of erythromycin-resistant Streptococcus pyogenes and correlations with macrolide-resistance phenotypes.

    PubMed

    Ripa, S; Zampaloni, C; Vitali, L A; Giovanetti, E; Montanari, M P; Prenna, M; Varaldo, P E

    2001-01-01

    High rates of erythromycin resistance among Streptococcus pyogenes strains have been reported in Italy in the last few years. In this study, 370 erythromycin-resistant (MIC, > or = 1 microg/mL) Italian isolates of this species obtained in 1997-1998 from throat swabs from symptomatic patients were typed by analyzing SmaI macrorestriction fragment patterns by pulsed-field gel electrophoresis (PFGE). Among the typable isolates (n = 341; the genomic DNA of the remaining 29 isolates was not restricted by SmaI), 48 distinct PFGE types were recognized, of which 31 were recorded in only one isolate (one-strain types). Fifty-two percent of typable isolates fell into three type clusters and 75% into six, suggesting that erythromycin-resistant group A streptococci circulating in Italy are polyclonal, but the majority of them probably derives from the spread of a limited number of clones. In parallel experiments, the 370 test strains were characterized for the macrolide resistance phenotype: 80 were assigned to phenotype cMLS, 89 to phenotype iMLS-A, 33 to phenotype iMLS-B, 11 to phenotype iMLS-C, and 157 to phenotype M. There was a close correlation between these phenotypic data and the genotypic results of PFGE analysis, the vast majority of the isolates assigned to individual PFGE classes belonging usually to a single phenotype of macrolide resistance. All of the 29 untypable isolates belonged to the M phenotype. Further correlations were observed with tetracycline resistance.

  14. A method for allelic replacement that uses the conjugative transposon Tn916: deletion of the emm6.1 allele in Streptococcus pyogenes JRS4.

    PubMed

    Norgren, M; Caparon, M G; Scott, J R

    1989-12-01

    The emm6.1 allele of Streptococcus pyogenes JRS4 was deleted by using the conjugative transposon Tn916. The aphA-3 gene, conferring resistance to kanamycin, was cloned between the sequences flanking the structural gene for the type 6 M protein (emm6.1) and inserted into the BstXI site of Tn916 to generate the chimeric transposon Tn916-5K3. Because the BstXI site lies in a nonessential region of Tn916, the chimeric transposon could transfer by conjugation from Bacillus subtilis into JRS4. In some of the transconjugants, Tn916-5K3 replaced the emm6.1 locus of JRS4 by homologous recombination between the cloned emm6.1-flanking regions and the resident chromosome. One recombinant studied in detail, JRS75, was kanamycin resistant and tetracycline sensitive and lacked immunologically detectable M6 protein. Furthermore, by Southern blot analysis, the DNA region encompassing the emm6.1 structural gene was found to have been replaced by aphA-3.

  15. Use of a Phosphorylation Site Mutant To Identify Distinct Modes of Gene Repression by the Control of Virulence Regulator (CovR) in Streptococcus pyogenes.

    PubMed

    Horstmann, Nicola; Sahasrabhojane, Pranoti; Yao, Hui; Su, Xiaoping; Shelburne, Samuel A

    2017-09-15

    Control of the virulence regulator/sensor kinase (CovRS) two-component system (TCS) serves as a model for investigating the impact of signaling pathways on the pathogenesis of Gram-positive bacteria. However, the molecular mechanisms by which CovR, an OmpR/PhoB family response regulator, controls virulence gene expression are poorly defined, partly due to the labile nature of its aspartate phosphorylation site. To better understand the regulatory effect of phosphorylated CovR, we generated the phosphorylation site mutant strain 10870-CovR-D53E, which we predicted to have a constitutive CovR phosphorylation phenotype. Interestingly, this strain showed CovR activity only for a subset of the CovR regulon, which allowed for classification of CovR-influenced genes into D53E-regulated and D53E-nonregulated groups. Inspection of the promoter sequences of genes belonging to each group revealed distinct promoter architectures with respect to the location and number of putative CovR-binding sites. Electrophoretic mobility shift analysis demonstrated that recombinant CovR-D53E protein retains its ability to bind promoter DNA from both CovR-D53E-regulated and -nonregulated groups, implying that factors other than mere DNA binding are crucial for gene regulation. In fact, we found that CovR-D53E is incapable of dimerization, a process thought to be critical to OmpR/PhoB family regulator function. Thus, our global analysis of CovR-D53E indicates dimerization-dependent and dimerization-independent modes of CovR-mediated repression, thereby establishing distinct mechanisms by which this critical regulator coordinates virulence gene expression.IMPORTANCEStreptococcus pyogenes causes a wide variety of diseases, ranging from superficial skin and throat infections to life-threatening invasive infections. To establish these various disease manifestations, Streptococcus pyogenes requires tightly coordinated production of its virulence factor repertoire. Here, the response regulator Cov

  16. Crystal structure of peroxide stress regulator from Streptococcus pyogenes provides functional insights into the mechanism of oxidative stress sensing.

    PubMed

    Makthal, Nishanth; Rastegari, Sheila; Sanson, Misu; Ma, Zhen; Olsen, Randall J; Helmann, John D; Musser, James M; Kumaraswami, Muthiah

    2013-06-21

    Regulation of oxidative stress responses by the peroxide stress regulator (PerR) is critical for the in vivo fitness and virulence of group A Streptococcus. To elucidate the molecular mechanism of DNA binding, peroxide sensing, and gene regulation by PerR, we performed biochemical and structural characterization of PerR. Sequence-specific DNA binding by PerR does not require regulatory metal occupancy. However, metal binding promotes higher affinity PerR-DNA interactions. PerR metallated with iron directly senses peroxide stress and dissociates from operator sequences. The crystal structure revealed that PerR exists as a homodimer with two metal-binding sites per subunit as follows: a structural zinc site and a regulatory metal site that is occupied in the crystals by nickel. The regulatory metal-binding site in PerR involves a previously unobserved HXH motif located in its unique N-terminal extension. Mutational analysis of the regulatory site showed that the PerR metal ligands are involved in regulatory metal binding, and integrity of this site is critical for group A Streptococcus virulence. Interestingly, the metal-binding HXH motif is not present in the structurally characterized members of ferric uptake regulator (Fur) family but is fully conserved among PerR from the genus Streptococcus. Thus, it is likely that the PerR orthologs from streptococci share a common mechanism of metal binding, peroxide sensing, and gene regulation that is different from that of well characterized PerR from Bacillus subtilis. Together, our findings provide key insights into the peroxide sensing and regulation of the oxidative stress-adaptive responses by the streptococcal subfamily of PerR.

  17. Crystal Structure of Peroxide Stress Regulator from Streptococcus pyogenes Provides Functional Insights into the Mechanism of Oxidative Stress Sensing*

    PubMed Central

    Makthal, Nishanth; Rastegari, Sheila; Sanson, Misu; Ma, Zhen; Olsen, Randall J.; Helmann, John D.; Musser, James M.; Kumaraswami, Muthiah

    2013-01-01

    Regulation of oxidative stress responses by the peroxide stress regulator (PerR) is critical for the in vivo fitness and virulence of group A Streptococcus. To elucidate the molecular mechanism of DNA binding, peroxide sensing, and gene regulation by PerR, we performed biochemical and structural characterization of PerR. Sequence-specific DNA binding by PerR does not require regulatory metal occupancy. However, metal binding promotes higher affinity PerR-DNA interactions. PerR metallated with iron directly senses peroxide stress and dissociates from operator sequences. The crystal structure revealed that PerR exists as a homodimer with two metal-binding sites per subunit as follows: a structural zinc site and a regulatory metal site that is occupied in the crystals by nickel. The regulatory metal-binding site in PerR involves a previously unobserved HXH motif located in its unique N-terminal extension. Mutational analysis of the regulatory site showed that the PerR metal ligands are involved in regulatory metal binding, and integrity of this site is critical for group A Streptococcus virulence. Interestingly, the metal-binding HXH motif is not present in the structurally characterized members of ferric uptake regulator (Fur) family but is fully conserved among PerR from the genus Streptococcus. Thus, it is likely that the PerR orthologs from streptococci share a common mechanism of metal binding, peroxide sensing, and gene regulation that is different from that of well characterized PerR from Bacillus subtilis. Together, our findings provide key insights into the peroxide sensing and regulation of the oxidative stress-adaptive responses by the streptococcal subfamily of PerR. PMID:23645680

  18. Effects of penicillin and erythromycin on adherence of invasive and noninvasive isolates of Streptococcus pyogenes to laminin

    PubMed Central

    Šmitran, Aleksandra; Vuković, Dragana; Gajić, Ina; Marinković, Jelena; Ranin, Lazar

    2015-01-01

    This study investigated the possible relationship between the invasiveness of group A Streptococcus (GAS) strains and their abilities to adhere to laminin and assessed the effects of subinhibitory concentrations of penicillin and erythromycin on the ability of GAS to adhere to laminin. The adherence of noninvasive and highly invasive isolates of GAS to laminin was significantly higher than the adherence displayed by isolates of low invasiveness. Antibiotic treatment caused significant reductions in adherence to laminin in all three groups of strains. Penicillin was more successful in reducing the adherence abilities of the tested GAS strains than erythromycin. PMID:26270594

  19. Preliminary pediatric clinical evaluation of the oral probiotic Streptococcus salivarius K12 in preventing recurrent pharyngitis and/or tonsillitis caused by Streptococcus pyogenes and recurrent acute otitis media.

    PubMed

    Di Pierro, Francesco; Donato, Guido; Fomia, Federico; Adami, Teresa; Careddu, Domenico; Cassandro, Claudia; Albera, Roberto

    2012-01-01

    The oral probiotic Streptococcus salivarius K12 has been shown clearly to antagonize the growth of Streptococcus pyogenes, the most important bacterial cause of pharyngeal infections in humans, by releasing two bacteriocins named salivaricin A2 and salivaricin B. Unpublished observations indicate that it can also antagonize the growth of other bacteria involved in acute otitis media. Because of its ability to colonize the oral cavity and its safety profile, we have tested its efficacy in reducing the incidence of streptococcal pharyngitis and/or tonsillitis and episodes of acute otitis media. We enrolled 82 children, including 65 with and 17 without a recent diagnosis of recurrent oral streptococcal pathology. Of those with recurrent pathology, 45 were treated daily for 90 days with an oral slow-release tablet containing five billion colony-forming units of S. salivarius K12 (Bactoblis(®)), and the remaining 20 served as an untreated control group. The 17 children without a recent diagnosis of recurrent oral pathology were used as an additional control group. After 90 days of treatment, a 6-month follow-up period without treatment was included to evaluate a possible persistent protective role for the previously administered product. The 41 children who completed the 90-day course of Bactoblis showed a reduction in their episodes of streptococcal pharyngeal infection (about 90%) and/or acute otitis media (about 40%), calculated by comparing infection rates in the previous year. The 90-day treatment also reduced the reported incidence of pharyngeal and ear infections by about 65% in the 6-month follow-up period during which the product was not administered. Subjects tolerated the product well, with no side effects or dropouts reported. Prophylactic administration of S. salivarius K12 to children with a history of recurrent oral streptococcal pathology reduced episodes of streptococcal pharyngeal infections and/or tonsillitis as well as episodes of acute otitis media.

  20. In vitro antimicrobial activity of ozenoxacin against methicillin-susceptible Staphylococcus aureus, methicillin-resistant S. aureus and Streptococcus pyogenes isolated from clinical cutaneous specimens in Japan.

    PubMed

    Kanayama, Shoji; Ikeda, Fumiaki; Okamoto, Kazuaki; Nakajima, Akiko; Matsumoto, Tatsumi; Ishii, Ritsuko; Amano, Ayako; Matsuzaki, Kaoru; Matsumoto, Satoru

    2016-10-01

    Ozenoxacin, a novel non-fluorinated topical quinolone, was assessed for in vitro antimicrobial activity against each 50 isolates of methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant S. aureus (MRSA), and Streptococcus pyogenes according to the broth microdilution method recommended by the Clinical and Laboratory Standards Institute. The isolates used in this study were recovered from cutaneous specimens of Japanese adult and pediatric patients who visited hospitals in 2014. The MIC90s of ozenoxacin against MSSA, MRSA and S. pyogenes isolates from adult patients were ≤0.06, 4 and ≤0.06 μg/mL, respectively. The MIC90s of ozenoxacin against MSSA and S. pyogenes isolates from pediatric patients were equal to those against the adult isolates. On the other hand, the MIC90s of ozenoxacin against the pediatric MRSA isolates was 0.12 μg/mL, and was 32 times lower than that against the adult isolates. The antimicrobial activity of ozenoxacin against MSSA, MRSA and S. pyogenes was equal to or greater than those of 7 reference antimicrobial agents had been used for the treatment of skin infections. The MICs of ozenoxacin was highly correlated with those of nadifloxacin and levofloxacin in the 50 MRSA isolates (r(2) = 0.906 and 0.833, respectively). However, ozenoxacin kept the potent antimicrobial activity with the MIC ranging from 1 to 4 μg/mL even against MRSA low susceptible (MIC: >64 μg/mL) to nadifloxacin or levofloxacin. Ozenoxacin could represent the first-in-class non-fluorinated quinolone for the topical treatment of various superficial skin infections caused by MSSA, MRSA and S. pyogenes.

  1. In Vitro Activity of the New Ketolide Telithromycin Compared with Those of Macrolides against Streptococcus pyogenes: Influences of Resistance Mechanisms and Methodological Factors

    PubMed Central

    Bemer-Melchior, Pascale; Juvin, Marie-Emmanuelle; Tassin, Sandrine; Bryskier, Andre; Schito, Gian Carlo; Drugeon, Henri-B.

    2000-01-01

    One hundred and seven clinical isolates of Streptococcus pyogenes, 80 susceptible to macrolides and 27 resistant to erythromycin A (MIC >0.5 μg/ml), were examined. The erythromycin A-lincomycin double-disk test assigned 7 resistant strains to the M-phenotype, 8 to the inducible macrolide, lincosamide, and streptogramin B resistance (iMLSB) phenotype, and 12 to the constitutive MLSB resistance (cMLSB) phenotype. MICs of erythromycin A, clarithromycin, azithromycin, roxithromycin, and clindamycin were determined by a broth microdilution method. MICs of telithromycin were determined by three different methods (broth microdilution, agar dilution, and E-test methods) in an ambient air atmosphere and in a 5 to 6% CO2 atmosphere. Erythromycin A resistance genes were investigated by PCR in the 27 erythromycin A-resistant isolates. MICs of erythromycin A and clindamycin showed six groups of resistant strains, groups A to F. iMLSB strains (A, B, and D groups) are characterized by two distinct patterns of resistance correlated with genotypic results. A- and B-group strains were moderately resistant to 14- and 15-membered ring macrolides and highly susceptible to telithromycin. All A- and B-group isolates harbored erm TR gene, D-group strains, highly resistant to macrolides and intermediately resistant to telithromycin (MICs, 1 to 16 μg/ml), were all characterized by having the ermB gene. All M-phenotype isolates (C group), resistant to 14- and 15-membered ring macrolides and susceptible to clindamycin and telithromycin, harbored the mefA gene. All cMLSB strains (E and F groups) with high level of resistance to macrolides, lincosamide, and telithromycin had the ermB gene. The effect of 5 to 6% CO2 was remarkable on resistant strains, by increasing MICs of telithromycin from 1 to 6 twofold dilutions against D-E- and F-group isolates. PMID:11036012

  2. Typing of the pilus-protein-encoding FCT region and biofilm formation as novel parameters in epidemiological investigations of Streptococcus pyogenes isolates from various infection sites.

    PubMed

    Köller, Thomas; Manetti, Andrea Guido Oreste; Kreikemeyer, Bernd; Lembke, Cordula; Margarit, Immaculada; Grandi, Guido; Podbielski, Andreas

    2010-04-01

    Streptococcus pyogenes is an important human pathogen for which an association between infection site and selected epidemiological or functional markers has previously been suggested. However, the studies involved often used strains with an insufficiently defined clinical background and laboratory history. Thus, the major goal of the present study was to investigate these relationships in 183 prospectively collected, well-defined, low-passage isolates from a North-East German centre for tertiary care. For each isolate the clinical background (91 respiratory, 71 skin and 21 invasive isolates) and antibiotic-resistance pattern was recorded. All isolates were classified according to their emm type, antibiotic-resistance and PFGE pattern ( SmaI restriction analysis of genomic DNA). As novel discriminatory methods we performed a PCR-based typing of the pilus-protein-encoding FCT region (FCT) and biofilm-formation phenotyping in various culture media. Forty-one isolates were found to be resistant to at least one of the tested antibiotics. emm typing revealed emm28, emm12 , emm1, emm4, emm89 and emm2 as the most frequent types in our collection. The novel FCT typing showed isolates encoding FCT types 4 and 2 to be the most common. Overall 113 strains with unique combinations of emm and FCT types, antibiotic-resistance and PFGE patterns were identified. The majority of all isolates revealed an association of biofilm-formation capacity with growth media. Comparing all results for potential associations, no correlation could be established between the anatomical site of isolation and the emm or the FCT type. There was no relationship between biofilm formation and emm type, antibiotic-resistance or PFGE patterns. However, a novel association between biofilm formation and FCT type became obvious among strains from our collection.

  3. Label-free proteomic analysis of environmental acidification-influenced Streptococcus pyogenes secretome reveals a novel acid-induced protein histidine triad protein A (HtpA) involved in necrotizing fasciitis.

    PubMed

    Wen, Yao-Tseng; Wang, Jie-Siou; Tsai, Shu-Han; Chuan, Chiang-Ni; Wu, Jiunn-Jong; Liao, Pao-Chi

    2014-09-23

    Streptococcus pyogenes is responsible for various diseases. During infection, bacteria must adapt to adverse environments, such as the acidic environment. Acidic stimuli may stimulate S. pyogenes to invade into deeper tissue. However, how this acidic stimulus causes S. pyogenes to manipulate its secretome for facilitating invasion remains unclear. The dynamic label-free LC-MS/MS profiling identified 97 proteins, which are influenced by environmental acidification. Among these, 33 (34%) of the identified proteins were predicted to be extracellular proteins. Interestingly, classical secretory proteins comprise approximately 90% of protein abundance of the secretome in acidic condition at the stationary phase. One acid-induced secreted protein, HtpA, was selected to investigate its role in invasive infection. The mouse infected by the htpA deficient mutant showed lower virulence and smaller lesion area than the wild-type strain. The mutant strain was more efficiently cleared at infected skin than the wild-type strain. Besides, the relative phagocytosis resistance is lower in the mutant strain than in the wild-type strain. These data indicate that a novel acid-induced virulence factor, HtpA, which improves anti-phagocytosis ability for causing necrotizing fasciitis. Our investigation provides vital information for documenting the broad influences and mechanisms underlying the invasive behavior of S. pyogenes in an acidified environment. The acidified infected environment may facilitate S. pyogenes invasion from the mucosa to the deeper subepithelial tissue. The acid stimuli have been considered to affect the complex regulatory network of S. pyogenes for causing severe infections. Many of secreted virulence factors influenced by acidified environment may also play a crucial role in pathogenesis of invasive disease. To investigate temporal secretome changes under acidic environment, a comparative secretomics approach using label-free LC-MS/MS was undertaken to analyze

  4. [Prevalence of Streptococcus pyogenes and methicillin-resistant Staphylococcus aureus in the pharynx of healthy children in the town of Gardabaer].

    PubMed

    Magnúsdóttir, Björg Thurídur; Jónsson, Jón Steinar; Kristinsson, Karl G

    2008-06-01

    Treating S. pyogenes pharyngitis with antibiotics is recommended after confirming its presence using culture or rapid antigen tests. Limiting unnecessary antibiotics use is important in attempt to avoid rising resistance to drugs such as macrolides. Not all individuals carrying S. pyogenes are infected. To evaluate the carriage rate of S. pyogenes and methicillin resistant S. aureus (MRSA) among healthy children in the Reykjavík capital area. Cross-sectional study for the carriage of S. pyogenes and MRSA among healthy children in the town of Gardabaer. The study took place in March and April 2005. Throat cultures were collected from 270 asymptomatic healthy primary school students and cultured selectively for S. pyogenes and MRSA and tested for antimicrobial susceptibilities. Prevalence of S. pyogenes was found to be 22%. The proportion of carriers in 1st to 6th grade was 28%, compared with 11% in 7th to 10th grade students. The highest proportion was in 1st grade, 45%. The proportion S. pyogenes resistant to erythromycin was 17%, to tetracycline 13% and clindamycin 2%. All strains were susceptible to penicillin. No MRSA strains were found. The study reveals a high S. pyogenes carriage rate in primary school children in Garethabaer. Physicians should consider the prevalence of streptococcal carriage when diagnosing streptococcal pharyngitis in children, and only perform culture and/or antigen tests when clinically indicated.

  5. The Streptococcus pyogenes orphan protein tyrosine phosphatase, SP-PTP, possesses dual specificity and essential virulence regulatory functions.

    PubMed

    Kant, Sashi; Agarwal, Shivani; Pancholi, Preeti; Pancholi, Vijay

    2015-08-01

    Group A Streptococcus (GAS) is a human pathogen that causes high morbidity and mortality. GAS lacks a gene encoding tyrosine kinase but contains one encoding tyrosine phosphatase (SP-PTP). Thus, GAS is thought to lack tyrosine phosphorylation, and the physiological significance of SP-PTP is, therefore, questionable. Here, we demonstrate that SP-PTP possesses dual phosphatase specificity for Tyr- and Ser/Thr-phosphorylated GAS proteins, such as Ser/Thr kinase (SP-STK) and the SP-STK-phosphorylated CovR and WalR proteins. Phenotypic analysis of GAS mutants lacking SP-PTP revealed that the phosphatase activity per se positively regulates growth, cell division and the ability to adhere to and invade host cells. Furthermore, A549 human lung cells infected with GAS mutants lacking SP-PTP displayed increased Ser-/Thr-/Tyr-phosphorylation. SP-PTP also differentially regulates the expression of ∼50% of the total GAS genes, including several virulence genes potentially through the two-component regulators, CovR, WalR and PTS/HPr regulation of Mga. Although these mutants exhibit attenuated virulence, a GAS mutant overexpressing SP-PTP is hypervirulent. Our study provides the first definitive evidence for the presence and importance of Tyr-phosphorylation in GAS and the relevance of SP-PTP as an important therapeutic target.

  6. Interleukin-17A Contributes to the Control of Streptococcus pyogenes Colonization and Inflammation of the Female Genital Tract

    PubMed Central

    Carey, Alison J.; Weinberg, Jason B.; Dawid, Suzanne R.; Venturini, Carola; Lam, Alfred K.; Nizet, Victor; Caparon, Michael G.; Walker, Mark J.; Watson, Michael E.; Ulett, Glen C.

    2016-01-01

    Postpartum women are at increased risk of developing puerperal sepsis caused by group A Streptococcus (GAS). Specific GAS serotypes, including M1 and M28, are more commonly associated with puerperal sepsis. However, the mechanisms of GAS genital tract infection are not well understood. We utilized a murine genital tract carriage model to demonstrate that M1 and M28 GAS colonization triggers TNF-α, IL-1β, and IL-17A production in the female genital tract. GAS-induced IL-17A significantly influences streptococcal carriage and alters local inflammatory responses in two genetically distinct inbred strains of mice. An absence of IL-17A or the IL-1 receptor was associated with reduced neutrophil recruitment to the site of infection; and clearance of GAS was significantly attenuated in IL-17A−/− mice and Rag1−/− mice (that lack mature lymphocytes) but not in mice deficient for the IL-1 receptor. Together, these findings support a role for IL-17A in contributing to the control of streptococcal mucosal colonization and provide new insight into the inflammatory mediators regulating host-pathogen interactions in the female genital tract. PMID:27241677

  7. Streptococcus pyogenes biofilm growth in vitro and in vivo and its role in colonization, virulence, and genetic exchange.

    PubMed

    Marks, Laura R; Mashburn-Warren, Lauren; Federle, Michael J; Hakansson, Anders P

    2014-07-01

    Group A streptococcus (GAS) commonly colonizes the oropharynx and nonintact skin. However, colonization has been little studied and the role of biofilm formation is unclear, as biofilm experiments to date have not been conducted under conditions that mimic the host environment. In this study we grew GAS biofilms on human keratinocytes under various environmental conditions and used this model to evaluate colonization, invasive disease and natural transformation. GAS grown on epithelial cells, but not biofilms grown on abiotic surfaces, produced biofilms with characteristics similar to in vivo colonization. These biofilm bacteria showed a 100-fold higher bacterial burden of nasal-associated lymphoid tissue in mice than broth-grown bacteria, and were not virulent during septic infection, which was attributed in part to down-regulation of genes typically involved in localized and invasive disease. We also showed for the first time that GAS were naturally transformable when grown in biofilms and during colonization of NALT in vivo. These findings provide novel model systems to study biofilm formation of GAS in vitro and in vivo, suggest an important role for biofilm formation during GAS colonization, and provide an explanation for the known genome diversity within the GAS population.

  8. Structural and functional analysis of RopB: A major virulence regulator in Streptococcus pyogenes

    SciTech Connect

    Makthal, Nishanth; Gavagan, Maire; Do, Hackwon; Olsen, Randall J.; Musser, James M.; Kumaraswami, Muthiah

    2016-02-19

    Group A Streptococcus (GAS) is an exclusive human pathogen that causes significant disease burden. Global regulator RopB of GAS controls the expression of several major virulence factors including secreted protease SpeB during high cell density. However, the molecular mechanism for RopB-dependent speB expression remains unclear. To understand the mechanism of transcription activation by RopB, we determined the crystal structure of the C-terminal domain of RopB. RopB-CTD has the TPR motif, a signature motif involved in protein-peptide interactions and shares significant structural homology with the quorum sensing RRNPP family regulators. Characterization of the high cell density-specific cell-free growth medium demonstrated the presence of a low molecular weight proteinaceous secreted factor that upregulates RopB-dependent speB expression. Together, these results suggest that RopB and its cognate peptide signals constitute an intercellular signalling machinery that controls the virulence gene expression in concert with population density. Structure-guided mutational analyses of RopB dimer interface demonstrated that single alanine substitutions at this critical interface significantly altered RopB-dependent speB expression and attenuated GAS virulence. Finally, results presented here suggested that a properly aligned RopB dimer interface is important for GAS pathogenesis and highlighted the dimerization interactions as a plausible therapeutic target for the development of novel antimicrobials.

  9. Serine/Threonine Phosphatase (SP-STP), Secreted from Streptococcus pyogenes, Is a Pro-apoptotic Protein*

    PubMed Central

    Agarwal, Shivani; Agarwal, Shivangi; Jin, Hong; Pancholi, Preeti; Pancholi, Vijay

    2012-01-01

    This investigation illustrates an important property of eukaryote-type serine/threonine phosphatase (SP-STP) of group A Streptococcus (GAS) in causing programmed cell death of human pharyngeal cells. The secretory nature of SP-STP, its elevated expression in the intracellular GAS, and the ability of wild-type GAS but not the GAS mutant devoid of SP-STP to cause apoptosis of the host cell both in vitro and in vivo suggest that GAS deploys SP-STP as an important virulence determinant to exploit host cell machinery for its own advantage during infection. The exogenously added SP-STP is able to enter the cytoplasm and subsequently traverses into the nucleus in a temporal fashion to cause apoptosis of the pharyngeal cells. The programmed cell death induced by SP-STP, which requires active transcription and de novo protein synthesis, is also caspase-dependent. Furthermore, the entry of SP-STP into the cytoplasm is dependent on its secondary structure as the catalytically inactive SP-STP with an altered structure is unable to internalize and cause apoptosis. The ectopically expressed wild-type SP-STP was found to be in the nucleus and conferred apoptosis of Detroit 562 pharyngeal cells. However, the catalytically inactive SP-STP was unable to cause apoptosis even when intracellularly expressed. The ability of SP-STP to activate pro-apoptotic signaling cascades both in the cytoplasm and in the nucleus resulted in mitochondrial dysfunctioning and perturbation in the phosphorylation status of histones in the nucleus. SP-STP thus not only functions as a virulence regulator but also as an important factor responsible for host-related pathogenesis. PMID:22262847

  10. Serine/threonine phosphatase (SP-STP), secreted from Streptococcus pyogenes, is a pro-apoptotic protein.

    PubMed

    Agarwal, Shivani; Agarwal, Shivangi; Jin, Hong; Pancholi, Preeti; Pancholi, Vijay

    2012-03-16

    This investigation illustrates an important property of eukaryote-type serine/threonine phosphatase (SP-STP) of group A Streptococcus (GAS) in causing programmed cell death of human pharyngeal cells. The secretory nature of SP-STP, its elevated expression in the intracellular GAS, and the ability of wild-type GAS but not the GAS mutant devoid of SP-STP to cause apoptosis of the host cell both in vitro and in vivo suggest that GAS deploys SP-STP as an important virulence determinant to exploit host cell machinery for its own advantage during infection. The exogenously added SP-STP is able to enter the cytoplasm and subsequently traverses into the nucleus in a temporal fashion to cause apoptosis of the pharyngeal cells. The programmed cell death induced by SP-STP, which requires active transcription and de novo protein synthesis, is also caspase-dependent. Furthermore, the entry of SP-STP into the cytoplasm is dependent on its secondary structure as the catalytically inactive SP-STP with an altered structure is unable to internalize and cause apoptosis. The ectopically expressed wild-type SP-STP was found to be in the nucleus and conferred apoptosis of Detroit 562 pharyngeal cells. However, the catalytically inactive SP-STP was unable to cause apoptosis even when intracellularly expressed. The ability of SP-STP to activate pro-apoptotic signaling cascades both in the cytoplasm and in the nucleus resulted in mitochondrial dysfunctioning and perturbation in the phosphorylation status of histones in the nucleus. SP-STP thus not only functions as a virulence regulator but also as an important factor responsible for host-related pathogenesis.

  11. Streptococcus pyogenes emm Types and Clusters during a 7-Year Period (2007 to 2013) in Pharyngeal and Nonpharyngeal Pediatric Isolates.

    PubMed

    Koutouzi, F; Tsakris, A; Chatzichristou, P; Koutouzis, E; Daikos, G L; Kirikou, E; Petropoulou, N; Syriopoulou, V; Michos, A

    2015-07-01

    Group A streptococcus (GAS) is an important cause of morbidity and mortality worldwide. Surveillance of emm types has important implications, as it can provide baseline information for possible implementation of vaccination. A total of 1,349 GAS pediatric isolates were collected during a 7-year period (2007 to 2013); emm typing was completed for 1,282 pharyngeal (84%) or nonpharyngeal (16%) isolates, and emm clusters and temporal changes were analyzed. Thirty-five different emm types, including 14 subtypes, were identified. The most prevalent emm types identified were 1 (16.7%), 12 (13.6%), 77 (10.9%), 4 (10.8%), 28 (10.4%), 6 (6.8%), 3 (6.6%), and 89 (6.6%), accounting for 82.3% of total isolates. Rheumatogenic emm types comprised 16.3% of total isolates. The emm types 12, 4, and 77 were more prevalent among pharyngeal isolates, and the emm types 1, 89, 6, 75, and 11 were more prevalent among nonpharyngeal isolates. The emm types identified belonged to 13 emm clusters, and the 8 most prevalent clusters comprised 97% of all isolates. There were statistically significant decreases in the prevalence of emm types 12, 4, 5, and 61 and increases in the prevalence of emm types 89, 75, and 11, compared with the period 2001 to 2006. The proposed 30-valent GAS vaccine, which is currently in preclinical studies, encompasses 97.2% of the emm types detected in our study and 97.4% of the erythromycin-resistant strains. In addition, it includes 93.3% of the emm types involved in bacteremia. A much greater diversity of GAS emm types was identified in our area than described previously. Seasonal fluctuations and the introduction of new emm types were observed. Continuous surveillance of emm types is needed in order to evaluate the possible benefits of an M protein-based GAS vaccine.

  12. Is there any difference in pyogenic liver abscess caused by Streptococcus milleri and Klebsiella spp?: retrospective analysis over a 10-year period in a regional hospital.

    PubMed

    Law, Siu-Tong; Kong Li, Michael Kin

    2013-02-01

    To compare the clinical characteristics of patients with Streptococcus milleri (SM) and Klebsiella spp. associated pyogenic liver abscess (PLA). A retrospective study of patients with PLA due to SM and Klebsiella spp. was conducted. Clinical characteristics, laboratory and radiological features, management and outcomes were analyzed. From 2000 to 2009 inclusive, 21 and 140 patients had SM and Klebsiella spp. associated monomicrobial infected PLA, respectively. A higher incidence of active malignancy occurred in the SM group (14.3% vs. 3.6%, p < 0.03). The common clinical features of the patients were fever, chill and right upper quadrant pain. A longer duration (6.3 vs. 4.4 day, p = 0.04) of symptoms and a higher incidence of hepatomegaly (14.3% vs. 2.9%, p < 0.01) occurred in the SM group. Common laboratory and imaging abnormalities included: anemia, leukocytosis, high erythrocyte sedimentation rate and C-reactive protein, hypoalbuminemia, elevated total bilirubin and alanine aminotransferase, right hepatic lobe involvement, hypoechoic in ultrasonograpghy, rim enhancement and septal lobulation in computed tomography. The biliary tract disorder was the most common cause of the disease in the two groups. Patients with Klebsiella spp. associated PLA tended to have more complications: bacteremia (61.6% vs. 31.6%, p < 0.01) septic shock (33.6% vs. 19%, p = 0.11), disseminated intravascular coagulation (20.7% vs. 4.8%, p = 0.04), metastatic infections (10.7% vs. 0%, p = 0.06), acute renal and respiratory failure (5% vs. 0%, p = 0.14). However, both were effectively managed by the combination of antibiotics and image-guided aspiration with/without drainage, and their mortality rates were comparable to each other. Those patients with metastatic infection might need a longer duration (6.07 vs. 5.32 week, p = 0.144) of antibiotic therapy, which was due to the longer mean duration (3.85 vs. 2.86, p < 0.04) of an intravenous counterpart. SM associated PLA tends to have a

  13. Point-Counterpoint: A Nucleic Acid Amplification Test for Streptococcus pyogenes Should Replace Antigen Detection and Culture for Detection of Bacterial Pharyngitis.

    PubMed

    Pritt, Bobbi S; Patel, Robin; Kirn, Thomas J; Thomson, Richard B

    2016-10-01

    Nucleic acid amplification tests (NAATs) have frequently been the standard diagnostic approach when specific infectious agents are sought in a clinic specimen. They can be applied for specific agents such as S. pyogenes, or commercial multiplex NAATs for detection of a variety of pathogens in gastrointestinal, bloodstream, and respiratory infections may be used. NAATs are both rapid and sensitive. For many years, S. pyogenes testing algorithms used a rapid and specific group A streptococcal antigen test to screen throat specimens, followed, in some clinical settings, by a throat culture for S. pyogenes to increase the sensitivity of its detection. Now S. pyogenes NAATs are being used with increasing frequency. Given their accuracy, rapidity, and ease of use, should they replace antigen detection and culture for the detection of bacterial pharyngitis? Bobbi Pritt and Robin Patel of the Mayo Clinic, where S. pyogenes NAATs have been used for well over a decade with great success, will explain the advantages of this approach, while Richard (Tom) Thomson and Tom Kirn of the NorthShore University HealthSystem will discuss their concerns about this approach to diagnosing bacterial pharyngitis. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  14. Presence of the tet(O) gene in erythromycin- and tetracycline-resistant strains of Streptococcus pyogenes and linkage with either the mef(A) or the erm(A) gene.

    PubMed

    Giovanetti, Eleonora; Brenciani, Andrea; Lupidi, Remo; Roberts, Marilyn C; Varaldo, Pietro E

    2003-09-01

    Sixty-three recent Italian clinical isolates of Streptococcus pyogenes resistant to both erythromycin (MICs >or=1 microg/ml) and tetracycline (MICs >or= 8 microg/ml) were genotyped for macrolide and tetracycline resistance genes. We found 19 isolates carrying the mef(A) and the tet(O) genes; 25 isolates carrying the erm(A) and tet(O) genes; and 2 isolates carrying the erm(A), tet(M), and tet(O) genes. The resistance of all erm(A)-containing isolates was inducible, but the isolates could be divided into two groups on the basis of erythromycin MICs of either >128 or 1 to 4 microg/ml. The remaining 17 isolates included 15 isolates carrying the erm(B) gene and 2 isolates carrying both the erm(B) and the mef(A) genes, with all 17 carrying the tet(M) gene. Of these, 12 carried Tn916-Tn1545-like conjugative transposons. Conjugal transfer experiments demonstrated that the tet(O) gene moved with and without the erm(A) gene and with the mef(A) gene. These studies, together with the results of pulsed-field gel electrophoresis experiments and hybridization assays with DNA probes specific for the tet(O), erm(A), and mef(A) genes, suggested a linkage of tet(O) with either erm(A) or mef(A) in erythromycin- and tetracycline-resistant S. pyogenes isolates. By amplification and sequencing experiments, we detected the tet(O) gene ca. 5.5 kb upstream from the mef(A) gene. This is the first report demonstrating the presence of the tet(O) gene in S. pyogenes and showing that it may be linked with another gene and can be moved by conjugation from one chromosome to another.

  15. Virulence gene regulation by CvfA, a putative RNase: the CvfA-enolase complex in Streptococcus pyogenes links nutritional stress, growth-phase control, and virulence gene expression.

    PubMed

    Kang, Song Ok; Caparon, Michael G; Cho, Kyu Hong

    2010-06-01

    Streptococcus pyogenes, a multiple-auxotrophic human pathogen, regulates virulence gene expression according to nutritional availability during various stages in the infection process or in different infection sites. We discovered that CvfA influenced the expression of virulence genes according to growth phase and nutritional status. The influence of CvfA in C medium, rich in peptides and poor in carbohydrates, was most pronounced at the stationary phase. Under these conditions, up to 30% of the transcriptome exhibited altered expression; the levels of expression of multiple virulence genes were altered, including the genes encoding streptokinase, CAMP factor, streptolysin O, M protein (more abundant in the CvfA(-) mutant), SpeB, mitogenic factor, and streptolysin S (less abundant). The increase of carbohydrates or peptides in media restored the levels of expression of the virulence genes in the CvfA(-) mutant to wild-type levels (emm, ska, and cfa by carbohydrates; speB by peptides). Even though the regulation of gene expression dependent on nutritional stress is commonly linked to the stringent response, the levels of ppGpp were not altered by deletion of cvfA. Instead, CvfA interacted with enolase, implying that CvfA, a putative RNase, controls the transcript decay rates of virulence factors or their regulators according to nutritional status. The virulence of CvfA(-) mutants was highly attenuated in murine models, indicating that CvfA-mediated gene regulation is necessary for the pathogenesis of S. pyogenes. Taken together, the CvfA-enolase complex in S. pyogenes is involved in the regulation of virulence gene expression by controlling RNA degradation according to nutritional stress.

  16. Nucleotide sequence of conjugative prophage Φ1207.3 (formerly Tn1207.3) carrying the mef(A)/msr(D) genes for efflux resistance to macrolides in Streptococcus pyogenes

    PubMed Central

    Iannelli, Francesco; Santagati, Maria; Santoro, Francesco; Oggioni, Marco R.; Stefani, Stefania; Pozzi, Gianni

    2014-01-01

    Genetic element Φ1207.3 (formerly Tn1207.3) is a prophage of Streptococcus pyogenes which carries the macrolide efflux resistance genes mef(A)/msr(D) and is capable of conjugal transfer among streptococci. Complete nucleotide sequence showed that Φ1207.3 is 52,491 bp in length and contained 58 open reading frames (ORFs). A manual homology-based annotation with functional prediction of the hypothetical gene product was possible only for 34 out of 58 ORFs. Φ1207.3 codes for two different C-methylation systems, several phage structural genes, a lysis cassette (composed by a holin and a peptidoglycan hydrolase), and three site-specific resolvases of the serine recombinase family. PMID:25538698

  17. Nucleotide sequence of conjugative prophage Φ1207.3 (formerly Tn1207.3) carrying the mef(A)/msr(D) genes for efflux resistance to macrolides in Streptococcus pyogenes.

    PubMed

    Iannelli, Francesco; Santagati, Maria; Santoro, Francesco; Oggioni, Marco R; Stefani, Stefania; Pozzi, Gianni

    2014-01-01

    Genetic element Φ1207.3 (formerly Tn1207.3) is a prophage of Streptococcus pyogenes which carries the macrolide efflux resistance genes mef(A)/msr(D) and is capable of conjugal transfer among streptococci. Complete nucleotide sequence showed that Φ1207.3 is 52,491 bp in length and contained 58 open reading frames (ORFs). A manual homology-based annotation with functional prediction of the hypothetical gene product was possible only for 34 out of 58 ORFs. Φ1207.3 codes for two different C-methylation systems, several phage structural genes, a lysis cassette (composed by a holin and a peptidoglycan hydrolase), and three site-specific resolvases of the serine recombinase family.

  18. Cloning and sequence analysis of a gene encoding a 67-kilodalton myosin-cross-reactive antigen of Streptococcus pyogenes reveals its similarity with class II major histocompatibility antigens.

    PubMed Central

    Kil, K S; Cunningham, M W; Barnett, L A

    1994-01-01

    The group A streptococcal sequela acute rheumatic fever (ARF) has been associated with immunological cross-reactivity between streptococcal and heart proteins. To identify Streptococcus pyogenes genes that encode a myosin cross-reactive antigen(s) recognized by ARF sera, a genomic library from an emm deletion strain (T28/51/4) was screened with a single ARF serum. A positively identified lambda EMBL3 clone (T.2.18) produced a protein which reacted with myosin-specific antibodies affinity purified from individual ARF sera. The recombinant protein was initially estimated to be 60 kDa in size by sodium dodecyl sulfate-polyacrylamide gel electrophoresis; however, upon sequence analysis it had a molecular mass equivalent to 67 kDa. Sera from patients with streptococcal infections, acute glomerulonephritis, and ARF were reactive with the recombinant 67-kDa protein. However, individual sera from healthy persons were negative or demonstrated low levels of reactivity with the 67-kDa antigen. The gene encoding the 67-kDa myosin-cross-reactive antigen was subcloned, and its nucleotide sequence was determined by using a combined strategy of DNA sequencing of the cloned gene and N-terminal amino acid sequencing of the protein expressed in Escherichia coli. The amino-terminal sequence deduced from the nucleotide sequence of an open reading frame was identical to that determined from the 67-kDa protein expressed in E. coli. The gene encoded 590 amino acids with a calculated molecular weight of 67,381. No cleavable signal peptide was detected with the 67-kDa protein expressed in E. coli. The deduced amino acid sequence of the 67-kDa protein did not exhibit significant similarity to any known streptococcal proteins. However, it was found to be 19% identical and 62% similar over 151 amino acid residues to the beta chain of mouse major histocompatibility complex class II antigen (I-Au). Similar degrees of homology to the beta chains of other murine and human class II haplotypes were

  19. Intertrigo Caused by Streptococcus pyogenes.

    PubMed

    Chiriac, Anca; Murgu, Alina; Coroș, Marius Florin; Naznean, Adrian; Podoleanu, Cristian; Stolnicu, Simona

    2017-05-01

    Well-demarcated, beefy-red lesions of the skin folds, without satellite lesions, are the clinical hallmarks of intertrigo, frequently misdiagnosed especially in young children. We present 6 cases of streptococcal intertrigo to draw attention to this easily diagnosed and treated, but frequently overlooked, infection. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. In silico and in vitro studies of cinnamaldehyde and their derivatives against LuxS in Streptococcus pyogenes: effects on biofilm and virulence genes.

    PubMed

    Beema Shafreen, Raja Mohmed; Selvaraj, Chandrabose; Singh, Sanjeev Kumar; Karutha Pandian, Shunmugiah

    2014-02-01

    The LuxS-based signalling pathway has an important role in physiological and pathogenic functions that are capable of causing different infections. In the present study, cinnamaldehyde (CN) and their derivatives were evaluated for their inhibitory efficiency against LuxS by molecular modelling, docking, dynamics and free-energy calculations. Sequence and structure-similarity analysis of LuxS protein, five different amino acids were found to be highly conserved, of which GLY128 was identified as the key residue involved in the effective binding of the ligands. Quantum-polarized ligand docking protocol showed that 2nitro and 4nitro CN has a higher binding efficiency than CN, which very well corroborates with the in vitro studies. COMSTAT analysis for the microscopic images of the S. pyogenes biofilm showed that the ligands have antibiofilm potential. In addition, the results of quantitative polymerase chain reaction (qPCR) analysis revealed that the transcripts treated with the compounds showed decrease in luxS expression, which directly reflects with the reduction in expression of speB. No substantial effect was observed on the virulence regulator (srv) transcript. These results confirm that speB is controlled by the regulation of luxS. The decreased rate of S. pyogenes survival in the presence of these ligands envisaged the fact that the compounds could readily enhance opsonophagocytosis with the reduction of virulence factor secretion. Thus, the overall data supports the use of CN derivatives against quorum sensing-mediated infections caused by S. pyogenes.

  1. A novel double-tryptophan peptide pheromone is conserved in mutans and pyogenic Streptococci and Controls Competence in Streptococcus mutans via an Rgg regulator

    PubMed Central

    Mashburn-Warren, Lauren; Morrison, Donald A.; Federle, Michael J.

    2010-01-01

    Summary All streptococcal genomes encode the alternative sigma factor SigX and 21 SigX-dependent proteins required for genetic transformation, yet no pyogenic streptococci are known to develop competence. Resolving this paradox may depend on understanding the regulation of sigX. We report the identification of a regulatory circuit linked to the sigX genes of both mutans and pyogenic streptococci that uses a novel small, double-tryptophan-containing competence-inducing peptide (CIP) pheromone. In both groups, the CIP gene, which we designate comS, and sigX have identical, noncanonical promoters consisting of 9-bp inverted repeats separated from a −10 hexamer by 19 bp. comS is adjacent to a gene encoding a putative transcription factor of the Rgg family and is regulated by its product, which we designate ComR. Deletion of comR or comS in S. mutans abolished transformability, as did deletion of the oligopeptide permease subunit oppD, suggesting that CIP is imported. Providing S. mutans with synthetic fragments of CIP revealed that seven C-terminal residues, including the WW motif, cause robust induction of both sigX and the competent state. We propose that this circuit is the proximal regulator of sigX in S. mutans, and we infer that it controls competence in a parallel way in all pyogenic streptococci. PMID:20969646

  2. EndoS from Streptococcus pyogenes is hydrolyzed by the cysteine proteinase SpeB and requires glutamic acid 235 and tryptophans for IgG glycan-hydrolyzing activity

    PubMed Central

    Allhorn, Maria; Olsén, Arne; Collin, Mattias

    2008-01-01

    Background The endoglycosidase EndoS and the cysteine proteinase SpeB from the human pathogen Streptococcus pyogenes are functionally related in that they both hydrolyze IgG leading to impairment of opsonizing antibodies and thus enhance bacterial survival in human blood. In this study, we further investigated the relationship between EndoS and SpeB by examining their in vitro temporal production and stability and activity of EndoS. Furthermore, theoretical structure modeling of EndoS combined with site-directed mutagenesis and chemical blocking of amino acids was used to identify amino acids required for the IgG glycan-hydrolyzing activity of EndoS. Results We could show that during growth in vitro S. pyogenes secretes the IgG glycan-hydrolyzing endoglycosidase EndoS prior to the cysteine proteinase SpeB. Upon maturation SpeB hydrolyzes EndoS that then loses its IgG glycan-hydrolyzing activity. Sequence analysis and structural homology modeling of EndoS provided a basis for further analysis of the prerequisites for IgG glycan-hydrolysis. Site-directed mutagenesis and chemical modification of amino acids revealed that glutamic acid 235 is an essential catalytic residue, and that tryptophan residues, but not the abundant lysine or the single cysteine residues, are important for EndoS activity. Conclusion We present novel information about the amino acid requirements for IgG glycan-hydrolyzing activity of the immunomodulating enzyme EndoS. Furthermore, we show that the cysteine proteinase SpeB processes/degrades EndoS and thus emphasize the importance of the SpeB as a degrading/processing enzyme of proteins from the bacterium itself. PMID:18182097

  3. Cloning and expression in Escherichia coli of the streptolysin O determinant from Streptococcus pyogenes: characterization of the cloned streptolysin O determinant and demonstration of the absence of substantial homology with determinants of other thiol-activated toxins.

    PubMed Central

    Kehoe, M; Timmis, K N

    1984-01-01

    A gene bank of Streptococcus pyogenes Richards was constructed in Escherichia coli by using the bacteriophage replacement vector lambda L47.1, and hybrid phage expressing streptolysin O (SLO) were identified among the recombinants. DNA sequences encoding SLO were subcloned from an slo+ hybrid phage into a low-copy-number vector plasmid to yield an slo+ hybrid plasmid, pMK157. This plasmid contains 5.6 kilobase pairs of cloned streptococcal DNA sequences, is stable, and expresses SLO at easily detectable levels in E. coli. Transposon gamma delta insertion mutants and in vitro-generated deletion mutants of pMK157 were isolated and analyzed. This analysis showed that a single gene is sufficient for production of SLO in E. coli and allowed this slo gene to be mapped to within +/- 100 base pairs. Two forms of the slo gene product, with molecular weights of 68,000 and 61,000, were detected in E. coli minicells harboring slo+ plasmids and by immunoblotting of E. coli whole cells harboring slo+ plasmids. Southern blotting hybridization experiments with the cloned SLO DNA sequences as probes failed to demonstrate homology between the cloned SLO determinant and DNA isolated from bacteria expressing thiol-activated cytolysins related to SLO. Images PMID:6321351

  4. 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.

    PubMed

    Swe, Pearl M; Christian, Lindsay D; Lu, Hieng C; Sriprakash, Kadaba S; Fischer, Katja

    2017-03-01

    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. 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. 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

  5. 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

    PubMed Central

    Swe, Pearl M.; Christian, Lindsay D.; Lu, Hieng C.; Sriprakash, Kadaba S.

    2017-01-01

    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

  6. Mutations in the control of virulence sensor gene from Streptococcus pyogenes after infection in mice lead to clonal bacterial variants with altered gene regulatory activity and virulence.

    PubMed

    Mayfield, Jeffrey A; Liang, Zhong; Agrahari, Garima; Lee, Shaun W; Donahue, Deborah L; Ploplis, Victoria A; Castellino, Francis J

    2014-01-01

    The cluster of virulence sensor (CovS)/responder (CovR) two-component operon (CovRS) regulates ∼15% of the genes of the Group A Streptococcal pyogenes (GAS) genome. Bacterial clones containing inactivating mutations in the covS gene have been isolated from patients with virulent invasive diseases. We report herein an assessment of the nature and types of covS mutations that can occur in both virulent and nonvirulent GAS strains, and assess whether a nonvirulent GAS can attain enhanced virulence through this mechanism. A group of mice were infected with a globally-disseminated clonal M1T1 GAS (isolate 5448), containing wild-type (WT) CovRS (5448/CovR+S+), or less virulent engineered GAS strains, AP53/CovR+S+ and Manfredo M5/CovR+S+. SpeB negative GAS clones from wound sites and/or from bacteria disseminated to the spleen were isolated and the covS gene was subjected to DNA sequence analysis. Numerous examples of inactivating mutations were found in CovS in all regions of the gene. The mutations found included frame-shift insertions and deletions, and in-frame small and large deletions in the gene. Many of the mutations found resulted in early translation termination of CovS. Thus, the covS gene is a genomic mutagenic target that gives GAS enhanced virulence. In cases wherein CovS- was discovered, these clonal variants exhibited high lethality, further suggesting that randomly mutated covS genes occur during the course of infection, and lead to the development of a more invasive infection.

  7. Complement-mediated Opsonization of Invasive Group A Streptococcus pyogenes Strain AP53 Is Regulated by the Bacterial Two-component Cluster of Virulence Responder/Sensor (CovRS) System*

    PubMed Central

    Agrahari, Garima; Liang, Zhong; Mayfield, Jeffrey A.; Balsara, Rashna D.; Ploplis, Victoria A.; Castellino, Francis J.

    2013-01-01

    Group A Streptococcus pyogenes (GAS) strain AP53 is a primary isolate from a patient with necrotizing fasciitis. These AP53 cells contain an inactivating mutation in the sensor component of the cluster of virulence (cov) responder (R)/sensor (S) two-component gene regulatory system (covRS), which enhances the virulence of the primary strain, AP53/covR+S−. However, specific mechanisms by which the covRS system regulates the survival of GAS in humans are incomplete. Here, we show a key role for covRS in the regulation of opsonophagocytosis of AP53 by human neutrophils. AP53/covR+S− cells displayed potent binding of host complement inhibitors of C3 convertase, viz. Factor H (FH) and C4-binding protein (C4BP), which concomitantly led to minimal C3b deposition on AP53 cells, further showing that these plasma protein inhibitors are active on GAS cells. This resulted in weak killing of the bacteria by human neutrophils and a corresponding high death rate of mice after injection of these cells. After targeted allelic alteration of covS− to wild-type covS (covS+), a dramatic loss of FH and C4BP binding to the AP53/covR+S+ cells was observed. This resulted in elevated C3b deposition on AP53/covR+S+ cells, a high level of opsonophagocytosis by human neutrophils, and a very low death rate of mice infected with AP53/covR+S+. We show that covRS is a critical transcriptional regulator of genes directing AP53 killing by neutrophils and regulates the levels of the receptors for FH and C4BP, which we identify as the products of the fba and enn genes, respectively. PMID:23928307

  8. Increased pilus production conferred by a naturally occurring mutation alters host-pathogen interaction in favor of carriage in Streptococcus pyogenes.

    PubMed

    Flores, Anthony R; Olsen, Randall J; Cantu, Concepcion; Pallister, Kyler B; Guerra, Fermin E; Voyich, Jovanka M; Musser, James M

    2017-03-06

    Studies of the human pathogen group A Streptococcus (GAS) define the carrier phenotype as increased ability to adhere to and persist on epithelial surfaces and decreased ability to cause disease. We tested the hypothesis that a single amino acid change (Arg135Gly) in a highly conserved sensor kinase (LiaS) of a poorly defined GAS regulatory system contributes to a carrier phenotype through increased pilus production. When introduced into an emm serotype-matched invasive strain, the carrier allele (liaS(R135G)) recapitulated a carrier phenotype defined by increased ability to adhere to mucosal surfaces and decreased ability to cause disease. Gene transcript analyses revealed that the liaS mutation significantly altered transcription of the genes encoding pilus when in the presence of bacitracin. Elimination of pilus production in the isogenic carrier mutant decreased ability to colonize the mouse nasopharynx, adhere to and be internalized by cultured human epithelial cells, and restored a virulence phenotype in a mouse model of necrotizing fasciitis. We also observed significantly reduced survival of the isogenic carrier mutant compared to the parental invasive strain after exposure to human neutrophils. Elimination of pilus in the isogenic carrier mutant increased neutrophil survival to the parental invasive strain level. Together, our data demonstrate that the carrier mutation (liaS(R135G)) affects pilus expression. Our data suggest new mechanisms of pilus gene regulation in GAS and differs from the enhanced invasiveness associated with increased pilus production in other bacterial pathogens.

  9. Prevalence of M75 Streptococcus pyogenes Strains Harboring slaA Gene in Patients Affected by Pediatric Obstructive Sleep Apnea Syndrome in Central Italy

    PubMed Central

    Viciani, Elisa; Montagnani, Francesca; Tordini, Giacinta; Romano, Antonio; Salerni, Lorenzo; De Luca, Andrea; Ruggiero, Paolo; Manetti, Andrea G. O.

    2017-01-01

    Recently we reported an association between pediatric obstructive sleep apnea syndrome (OSAS) and Group A streptococcus (GAS) sub-acute chronic tonsil colonization. We showed that GAS may contribute to tonsil hyperplasia via a streptolysin O (SLO)-dependent cysteinyl leukotrienes (CysLTs) production, which can trigger T and B cell proliferation. In the present study, we characterized the GAS strains isolated from pediatric OSAS patients in comparison with a panel of age and sex matched GAS strains unrelated to OSAS, but isolated in the same area and during the same period ranging from 2009 to 2013. We found that slaA gene, previously reported to be associated to CysLTs production pathway, was significantly associated to GAS OSAS strains. Moreover, the most numerous group (32%) of the GAS OSAS strains belonged to M75 type, and 6 out of 7 of these strains harbored the slaA gene. Multilocus Sequence Typing (MLST) experiments demonstrated that the clone emm75/ST49/ smeZ, slaA was associated to OSAS cases. In conclusion, we found an association between slaA gene and the GAS OSAS strains, and we showed that the clone emm75/ST49 harboring genes smeZ and slaA was exclusively isolated from patients affected by OSAS, thus suggesting that this genotype might be associated to the pathogenesis of OSAS, although further studies are needed to elucidate the possible role of SlaA in tonsil hypertrophy development. PMID:28293224

  10. Nonpolar Inactivation of the Hypervariable Streptococcal Inhibitor of Complement Gene (sic) in Serotype M1 Streptococcus pyogenes Significantly Decreases Mouse Mucosal Colonization

    PubMed Central

    Lukomski, Slawomir; Hoe, Nancy P.; Abdi, Iman; Rurangirwa, Jacqueline; Kordari, Parichher; Liu, Mengyao; Dou, Shu-Jun; Adams, Gerald G.; Musser, James M.

    2000-01-01

    Group A Streptococcus (GAS) is a human pathogen that commonly infects the upper respiratory tract. GAS serotype M1 strains are frequently isolated from human infections and contain the gene encoding the hypervariable streptococcal inhibitor of complement protein (Sic). It was recently shown that Sic variants were rapidly selected on mucosal surfaces in epidemic waves caused by M1 strains, an observation suggesting that Sic participates in host-pathogen interactions on the mucosal surface (N. P. Hoe, K. Nakashima, S. Lukomski, D. Grigsby, M. Liu, P. Kordari, S.-J. Dou, X. Pan, J. Vuopio-Varkila, S. Salmelinna, A. McGeer, D. E. Low, B. Schwartz, A. Schuchat, S. Naidich, D. De Lorenzo, Y.-X. Fu, and J. M. Musser, Nat. Med. 5:924–929, 1999). To test this idea, a new nonpolar mutagenesis method employing a spectinomycin resistance cassette was used to inactivate the sic gene in an M1 GAS strain. The isogenic Sic-negative mutant strain was significantly (P < 0.019) impaired in ability to colonize the mouse mucosal surface after intranasal infection. These results support the hypothesis that the predominance of M1 strains in human infections is related, in part, to a Sic-mediated enhanced colonization ability. PMID:10639414

  11. Comparative pathogenomic characterization of a non-invasive serotype M71 strain Streptococcus pyogenes NS53 reveals incongruent phenotypic implications from distinct genotypic markers.

    PubMed

    Bao, Yun-Juan; Li, Yang; Liang, Zhong; Agrahari, Garima; Lee, Shaun W; Ploplis, Victoria A; Castellino, Francis J

    2017-07-31

    The strains serotyped as M71 from group A Streptococcus are common causes of pharyngeal and skin diseases worldwide. Here we characterize the genome of a unique non-invasive M71 human isolate, NS53. The genome does not contain structural rearrangements or large-scale gene gains/losses, but encodes a full set of non-truncated known virulence factors, thus providing an ideal reference for comparative studies. However, the NS53 genome showed incongruent phenotypic implications from distinct genotypic markers. NS53 is characterized as an emm pattern D and FCT (fibronectin-collagen-T antigen) type-3 strain, typical of skin tropic strains, but is phylogenetically close to emm pattern E strains with preference for both skin and pharyngeal infections. We propose that this incongruence could result from recombination within the emm gene locus, or, alternatively, selection has been against those genetic alterations. Combined with the inability to select for CovS switching, a process is indicated whereby NS53 has been pre-adapted to specific host niches selecting against variations in CovS and many other genes. This may allow the strain to attain successful colonization and long-term survival. A balance between genetic variations and fitness may exist for this bacterium to form a stabilized genome optimized for survival in specific host environments. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  12. Molecular epidemiology, antimicrobial susceptibilities and resistance mechanisms of Streptococcus pyogenes isolates resistant to erythromycin and tetracycline in Spain (1994–2006)

    PubMed Central

    2012-01-01

    Background Group A Streptococcus (GAS) causes human diseases ranging in severity from uncomplicated pharyngitis to life-threatening necrotizing fasciitis and shows high rates of macrolide resistance in several countries. Our goal is to identify antimicrobial resistance in Spanish GAS isolates collected between 1994 and 2006 and to determine the molecular epidemiology (emm/T typing and PFGE) and resistance mechanisms of those resistant to erythromycin and tetracycline. Results Two hundred ninety-five out of 898 isolates (32.8%) were erythromycin resistant, with the predominance of emm4T4, emm75T25, and emm28T28, accounting the 67.1% of the 21 emm/T types. Spread of emm4T4, emm75T25 and emm28T28 resistant clones caused high rates of macrolide resistance. The distribution of the phenotypes was M (76.9%), cMLSB (20.3%), iMLSB (2.7%) with the involvement of the erythromycin resistance genes mef(A) (89.5%), msr(D) (81.7%), erm(B) (37.3%) and erm(A) (35.9%). Sixty-one isolates were tetracycline resistant, with the main representation of the emm77T28 among 20 emm/T types. To note, the combination of tet(M) and tet(O) tetracycline resistance genes were similar to tet(M) alone reaching values close to 40%. Resistance to both antibiotics was detected in 19 isolates of 7 emm/T types, being emm11T11 and the cMLSB phenotype the most frequent ones. erm(B) and tet(M) were present in almost all the strains, while erm(A), mef(A), msr(D) and tet(O) appeared in less than half of them. Conclusions Spanish GAS were highly resistant to macrolides meanwhile showed minor resistance rate to tetracycline. A remarkable correlation between antimicrobial resistance and emm/T type was noticed. Clonal spread of emm4T4, emm75T25 and emm28T28 was the main responsable for macrolide resistance where as that emm77T28 clones were it to tetraclycline resistance. A wide variety of macrolide resistance genes were responsible for three macrolide resistance phenotypes. PMID:22998619

  13. Effect of poly-hexamethylene biguanide hydrochloride (PHMB) treated non-sterile medical gloves upon the transmission of Streptococcus pyogenes, carbapenem-resistant E. coli, MRSA and Klebsiella pneumoniae from contact surfaces.

    PubMed

    Ali, S; Wilson, A P R

    2017-08-17

    Reduction of accidental contamination of the near-patient environment has potential to reduce acquisition of healthcare-associated infection(s). Although medical gloves should be removed when soiled or touching the environment, compliance is variable. The use of antimicrobial-impregnated medical gloves could reduce the horizontal-transfer of bacterial contamination between surfaces. Determine the activity of antimicrobial-impregnated gloves against common hospital pathogens: Streptococcus pyogenes, carbapenem-resistant E.coli (CREC), MRSA and ESBL-producing Klebsiella pneumoniae. Fingerpads (~1cm(2)) of PHMB-treated and untreated gloves were inoculated with 10 μL (~10(4) colony-forming-units [cfu]) of test-bacteria prepared in heavy-soiling (0.5%BSA), blood or distilled-water (no-soiling) and sampled after 0.25, 1, 10 or 15 min contact-time. Donor surfaces (~1cm(2) computer-keys) contaminated with wet/dry inoculum were touched with the fingerpad of treated/untreated gloves and subsequently pressed onto recipient (uncontaminated) computer-keys. Approximately 4.50log10cfu of all bacteria persisted after 15 min on untreated gloves regardless of soil-type. In the absence of soiling, PHMB-treated gloves reduced surface-contamination by ~4.5log10cfu (>99.99%) within 10 min of contact-time but only ~2.5log10 (>99.9%) and ~1.0log10 reduction respectively when heavy-soiling or blood was present. Gloves became highly-contaminated (~4.52log10-4.91log10cfu) when handling recently-contaminated computer-keys. Untreated gloves contaminated "recipient" surfaces (~4.5log10cfu) while PHMB-treated gloves transferred fewer bacteria (2.4-3.6log10cfu). When surface contamination was dry, PHMB gloves transferred fewer bacteria (0.3-0.6log10cfu) to "recipient" surfaces than untreated gloves (1.0-1.9log10; P < 0.05). Antimicrobial-impregnated gloves may be useful in preventing dissemination of organisms in the near-patient environment during routine care. However they are not a

  14. Pyogenic abscess (image)

    MedlinePlus

    ... become infected. The most common infecting bacteria include E coli , enterococcus, staphylococcus, and streptococcus. Treatment is usually a combination of drainage and prolonged antibiotic therapy.

  15. Pyogenic liver abscess

    PubMed Central

    Webb, Gwilym James; Chapman, Thomas Patrick; Cadman, Philip John; Gorard, David Angelo

    2014-01-01

    Pyogenic liver abscess has a variable clinical presentation. Its management requires input from several disciplines and is often coordinated by a gastroenterologist. This review examines demographics, clinical presentation, aetiology, diagnosis and prognosis; a suggested management approach, including antibiotic selection, radiological intervention and indications for surgery, is offered from a physician's perspective. PMID:28839753

  16. Pyogenic Brain Abscess in Thailand

    PubMed Central

    Wiwanitkit, Somsri; Wiwanitkit, Viroj

    2012-01-01

    Pyogenic brain abscess is important in neurology. This infectious disease is fatal and the management is usually complicated. Here, the authors review and discuss the clinical aspects of pyogenic brain abscess found in the earlier reports from a tropical setting in Thailand. The literature review was compiled through standard reference database searching and the derived publications were further extracted to obtain clinical data. The main clinical characteristics of pyogenic brain abscess in this setting were similar to others. However, there are some specific characteristics on the nature of tropical setting. PMID:22754873

  17. Primary pyogenic ventriculitis caused by Neisseria meningitidis: case report and review of the literature

    PubMed Central

    Hassan, Ibrahim; Newton, Pippa

    2017-01-01

    Background. Pyogenic ventriculitis is a well-known complication of meningitis, brain abscesses and intraventricular drains. Primary pyogenic ventriculitis is a rare entity and few cases have been described so far. We report the first case of primary pyogenic ventriculitis in an adult caused by Neisseria meningitidis and present an overview of all reported adult primary pyogenic ventriculitis cases in the English literature. Methods. A PubMed search was performed using the terms ependymitis, ventricular empyema, pyocephalus and ventriculitis. Filter was set for adults and English. Articles in which pyogenic ventriculitis was a complication of well-known risk factors were excluded. A total of five cases of primary pyogenic ventriculitis were identified. Results. There were seven adult patients. Only one patient showed signs of meningeal irritation. Four patients had positive blood cultures with Escherichia coli (one patient), methicillin-resistant Staphylococcus aureus (one patient), one patient was bacteraemic with Enterococcus faecalis, Escherichia coli and Peptostreptococcus spp., and N. meningitidis (our patient). In four patients cerebrospinal fluid was sent for culture, which yielded methicillin-sensitive Staphylococcus aureus (one patient), Peptostreptococcus spp. (one patient), Streptococcus intermedius (one patient, identified via 16S PCR) and Listeria monocytogenes (one patient). Cerebrospinal fluid cell count was determined in four patients and showed pleocytosis in all four cases. Ventricular drainage was performed in four patients. Five patients survived. Discussion. We report the first case of pyogenic ventriculitis caused by N. meningitidis. Primary pyogenic ventriculitis is a rare entity with various clinical presentations caused by various bacterial species. Treatment consists of adequate antimicrobial therapy, and ventricular drainage may be necessary. PMID:28348798

  18. Direct Host Plasminogen Binding to Bacterial Surface M-protein in Pattern D Strains of Streptococcus pyogenes Is Required for Activation by Its Natural Coinherited SK2b Protein.

    PubMed

    Chandrahas, Vishwanatha; Glinton, Kristofor; Liang, Zhong; Donahue, Deborah L; Ploplis, Victoria A; Castellino, Francis J

    2015-07-24

    Streptokinase (SK), secreted by Group A Streptococcus (GAS), is a single-chain ∼47-kDa protein containing three consecutive primary sequence regions that comprise its α, β, and γ modules. Phylogenetic analyses of the variable β-domain sequences from different GAS strains suggest that SKs can be arranged into two clusters, SK1 and SK2, with a subdivision of SK2 into SK2a and SK2b. SK2b is secreted by skin-tropic Pattern D M-protein strains that also express plasminogen (human Pg (hPg)) binding Group A streptococcal M-protein (PAM) as its major cell surface M-protein. SK2a-expressing strains are associated with nasopharynx tropicity, and many of these strains express human fibrinogen (hFg) binding Pattern A-C M-proteins, e.g. M1. PAM interacts with hPg directly, whereas M1 binds to hPg indirectly via M1-bound hFg. Subsequently, SK is secreted by GAS and activates hPg to plasmin (hPm), thus generating a proteolytic surface on GAS that enhances its dissemination. Due to these different modes of hPg/hPm recognition by GAS, full characterizations of the mechanisms of activation of hPg by SK2a and SK2b and their roles in GAS virulence are important topics. To more fully examine these subjects, isogenic chimeric SK- and M-protein-containing GAS strains were generated, and the virulence of these chimeric strains were analyzed in mice. We show that SK and M-protein alterations influenced the virulence of GAS and were associated with the different natures of hPg activation and hPm binding. These studies demonstrate that GAS virulence can be explained by disparate hPg activation by SK2a and SK2b coupled with the coinherited M-proteins of these strains. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. Streptococcus zooepidemicus and Streptococcus equi evolution: the role of CRISPRs.

    PubMed

    Waller, Andrew S; Robinson, Carl

    2013-12-01

    The host-restricted bacterium Streptococcus equi is the causative agent of equine strangles, the most frequently diagnosed infectious disease of horses worldwide. The disease is characterized by abscessation of the lymph nodes of the head and neck, leading to significant welfare and economic cost. S. equi is believed to have evolved from an ancestral strain of Streptococcus zooepidemicus, an opportunistic pathogen of horses and other animals. Comparison of the genome of S. equi strain 4047 with those of S. zooepidemicus identified examples of gene loss due to mutation and deletion, and gene gain through the acquisition of mobile genetic elements that have probably shaped the pathogenic specialization of S. equi. In particular, deletion of the CRISPR (clustered regularly interspaced short palindromic repeats) locus in the ancestor of S. equi may have predisposed the bacterium to acquire and incorporate new genetic material into its genome. These include four prophages and a novel integrative conjugative element. The virulence cargo carried by these mobile genetic elements is believed to have shaped the ability of S. equi to cause strangles. Further sequencing of S. zooepidemicus has highlighted the diversity of this opportunistic pathogen. Again, CRISPRs are postulated to influence evolution, balancing the need for gene gain over genome stability. Analysis of spacer sequences suggest that these pathogens may be susceptible to a limited range of phages and provide further evidence of cross-species exchange of genetic material among Streptococcus pyogenes, Streptococcus agalactiae and Streptococcus dysgalactiae.

  20. Streptococcus iniae and Streptococcus agalactiae

    USDA-ARS?s Scientific Manuscript database

    Streptococcus iniae and S. agalactiae are economically important Gram positive bacterial pathogens of cultured and wild fish with a worldwide distribution. Both bacteria are potential zoonotic pathogens and have been associated most often with infections in immunocompromised people. Streptococcus in...

  1. Pyogenic flexor tenosynovitis in children.

    PubMed

    Luria, Shai; Haze, Amir

    2011-08-01

    Pyogenic flexor tenosynovitis is an uncommon, emergent hand infection. The literature lacks any description of the disease and the variability of its manifestations in young children. We describe 3 cases. Two cases were diagnosed and treated promptly, and the third presented late, with atypical clinical signs, causing a delay in his diagnosis and treatment and stressing the caution to be taken with the evaluation of these children with signs of hand infection.

  2. Extragingival Pyogenic Granuloma: an Unusual Clinical Presentation

    PubMed Central

    Sachdeva, Suresh K.

    2015-01-01

    Pyogenic granuloma is thought to represent an exuberant tissue reaction to local irritation. It occurs in second decade of life in young females. Clinically, oral pyogenic granuloma is a smooth or lobulated exophytic growth, pedunculated or sessile, which usually bleeds on provocation. Oral pyogenic granuloma preferentially affects the gingiva. On rare occasion, it can be found extragingivally on lips, tongue, buccal mucosa, and palate which may mimic more serious pathological conditions such as malignancies. This article reports an unusual case of extra gingival pyogenic granuloma occurring on the right buccal mucosa in a female patient and discusses the features that distinguish this lesion from other similar oral mucosal lesions. PMID:26535410

  3. Nutritional requirements of Corynebacterium pyogenes.

    PubMed

    Fraga, A M; Reddy, C A

    1982-08-01

    The nutritional requirements of Corynebacterium pyogenes (strains C100, 5, and 1909), a commonly encountered animal pathogen, were determined in this study. A semidefined medium (SDM) containing glucose, HCO3-, hemin, charcoal-treated Trypticase, and a defined mixture of purines and pyrimidines, amino acids, and minerals which supported optimal growth of C. pyogenes was employed in all nutritional studies. Adenine and uracil were required for optimal growth of strains 5 and C100 but were not required for strain 1909. Riboflavin and nicotinic acid were required for good growth of all three strains; biotin and thiamin were stimulatory but did not appear to be required for growth. Hemin and NaHCO3 were stimulatory for growth, whereas lipoic acid and Tween 80 were neither stimulatory nor required for growth. The replacement of Trypticase with a specific peptide fraction (obtained by fractionation of Trypticase on Sephadex G-25) rich in dipeptides gave growth comparable to that in SDM, indicating a peptide requirement for the growth of C. pyogenes. It was of considerable interest that growth comparable to that in SDM was obtained when Trypticase was replaced by inositol (1 microgram/ml of SDM).

  4. Nutritional requirements of Corynebacterium pyogenes.

    PubMed Central

    Fraga, A M; Reddy, C A

    1982-01-01

    The nutritional requirements of Corynebacterium pyogenes (strains C100, 5, and 1909), a commonly encountered animal pathogen, were determined in this study. A semidefined medium (SDM) containing glucose, HCO3-, hemin, charcoal-treated Trypticase, and a defined mixture of purines and pyrimidines, amino acids, and minerals which supported optimal growth of C. pyogenes was employed in all nutritional studies. Adenine and uracil were required for optimal growth of strains 5 and C100 but were not required for strain 1909. Riboflavin and nicotinic acid were required for good growth of all three strains; biotin and thiamin were stimulatory but did not appear to be required for growth. Hemin and NaHCO3 were stimulatory for growth, whereas lipoic acid and Tween 80 were neither stimulatory nor required for growth. The replacement of Trypticase with a specific peptide fraction (obtained by fractionation of Trypticase on Sephadex G-25) rich in dipeptides gave growth comparable to that in SDM, indicating a peptide requirement for the growth of C. pyogenes. It was of considerable interest that growth comparable to that in SDM was obtained when Trypticase was replaced by inositol (1 microgram/ml of SDM). PMID:6288763

  5. Murine model of cutaneous infection with Streptococcus pyogenes.

    PubMed

    Medina, Eva

    2010-01-01

    Despite the medical advances achieved during the last century to fight against bacteria, viruses, fungi and parasites, infectious diseases are still a major cause of death, disability, and social and economic upheaval for millions around the world. Challenges remain in countering microorganisms even where antibiotics and vaccines are available. Much remains to be learned about basic aspects of the host-pathogen relationship and the complexity of the immune response to infection. Animal models represent a powerful tool to dissect the host response to infection, as well as the pathogenesis of the microbe. One of the advantages of using animal models is that both genetic and environmental factors that may influence the course of an infection can be controlled, allowing a precise cause-effect analysis of the host-pathogen interactions. In addition, there are no real alternatives to whole animal models in the study of integrative physiology and dynamic pathophysiologic alterations. The use of animal models has also proven invaluable for testing the efficacy of experimental antimicrobial agents and their therapeutic regimes. The mouse model is the most widely used for many reasons, including its cost effectiveness, the high number of immunological reagents available for this species, and the relative ease of biocontainment. Mouse strains with specific properties such as transgenic mouse strains with gene insertion or targeted mutation (knock-out) are very effective tools for studying the role of specific genes controlling the immune response to infectious pathogens. Murine models will remain the most appropriate tool for evaluating new the