Nizet Lab: Streptococci


	*Streptococci*

Basic
Microbiology Classification of
Streptococci Group A
Streptococcus Group B
Streptococcus

Basic Microbiology	Classification of Streptococci	Group A Streptococcus	Group B Streptococcus

Limited Time Special Offer!

Click on the graphic at right for a web-based version of the
P.I.'s lecture on Group A and Group B Streptococci,
delivered at the University of Minnesota (3/10/04):

Basic Microbiology
Dividing Strep The genus Streptococcus is made up of non-motile, Gram-positive cocci that grow in chains because when they divide each cross wall is made perpendicular to the long axis of the developing chain. Metabolically, streptococci are anaerobes because they obtain energy by fermentation, even when grown in air. Some are facultative anaerobes while others are strict anaerobes. Because they are not able to synthesize cytochromes, they do not carry out oxidative phosphorylation. The principle end product of fermentation is lactic acid. Therefore, streptococci are more acid-tolerant than most bacteria, which affects their ecology. Another consequence of lacking cytochromes is the absence of the cytochrome-dependent enzyme catalase (2 H2O2 --> 2 H 2O + O2). This feature is useful for identifying streptococci since most other bacteria make catalase; it also explains why streptococci are so susceptible to killing by polymorphonuclear leukocytes that make hydrogen peroxide as a product of the respiratory burst. Nearly all medically important streptococci have multiple nutritional requirements (auxotrophies) that include vitamins, amino acids, and nucleic acids, and are therefore not free-living in the environment. This also means that streptococci grow only in nutritionally rich media, and colonies are usually small.

Classification of Streptococci

Hemolysis The most common classification system for streptococci used everyday in the clinical microbiology lab is based on the appearance of colonies grown on the surface of blood-agar media. As depicted in the photograph to the left, streptococci are designated as exhibiting either alpha, beta or gamma hemolytic activity. In so-called ß-hemolysis , the RBCs surrounding the colony are completely lysed. Specific protein "hemolysin" toxins are responsible for this phenotype. The important pathogens in this group are group A streptococci and group B streptococci. In the case of alpha-hemolysis , the RBCs are intact but their pigment is changed from red to green. Hydrogen peroxide production is responsible for this phenotype, and these streptococci are sometimes referred to as "viridans" (= green). The most important human pathogen in this group is Streptococcus pneumoniae . Gamma-hemolysis is in fact a misnomer, as there is no change in the appearance of the RBC.

The ß-hemolytic streptococci are further classified on the basis of a scheme developed by Rebecca Lancefield (1895-1981) that tests the serologic reactivity of carbohydrate antigens (C substance) derived from acid extraction of the bacterial cell walls. Recognized serogroups are given letter designations from A-H and K-V. Some group antigens are shared by multiple species; however, only a single pathogenic species each comprises groups A (S. pyogenes) and B (S. agalactiae ). Other streptococci with pathogenic potential are found within the ß-hemolytic groups C, F and G (S. equisimilis or S. anginosus), but only extremely rarely is disease is associated with group D (S. bovis, S. durans or S. avium ) or other Lancefield groups. [Enterococcus faecalis and E. faecium were once classified as Group D streptococci]. S. pneumoniae (pneumococcus), a human pathogen producing pneumonia and meningitis, lacks a group specific antigen. Likewise, no group antigen is present in the viridans streptococci (e.g. S. mutans , S. sanguis, S. salivarius, S. milleri ) which are part of the oral flora and can contribute to the pathogenesis of dental caries or produce endocarditis on damaged heart valves.

Group A Streptococcus ( S. pyogenes , GAS)

Clinical Manifestations

GAS cause infections of the upper respiratory tract (principally pharyngitis = "strep throat", cervical lymphadenitis, peritonsillar abscess, pneumonia and empyema), and the skin (impetigo, lymphangitis, cellulitis and erysipelas). In addition, GAS are the cause of scarlet fever, and two immunologically mediated diseases, rheumatic fever and acute glomerulonephritis. Since the mid-1980's, a significant increase in deep and systemic infections caused by GAS has been observed, in particular necrotizing fasciitis and septicemia, often complicated by development of streptococcal toxic shock syndrome (STSS) and multiorgan system failure.

GAS Clinical Images

Watch a CNN News video clip describing invasive GAS infections including necrotizing fasciitis
Copyright: (c) 2000 Cable News Network, an AOL Time Warner Company. All rights reserved
(Requires the free RealPlayer software)

Epidemiology

GAS are exclusively human pathogens under natural conditions. Bacteria are passed from person to person, either by droplets, or by direct contact with infected skin, or possibly by fomites. Pharyngitis is more common in the winter and spring in temperate climates, and impetigo-like skin infections are more prevalent in the summer, often due to secondary infections of insect bites. The prevalence of GAS in the throats of school children varies from 5-40%. The highest incidence of GAS infections is in pre-adolescents. Skin trauma and chickenpox are important predisposing factors for severe, invasive GAS infections.

Pathogenesis

The pathogenesis of GAS infections begins with attachment to epithelial cells of the pharynx or skin via hairlike fimbriae that protrude from the cell surface and into the organism's hyaluronic acid capsule. The fimbriae are composed of lipotechoic acid and a series of serologically distinct surface proteins, the M proteins, which have important antiphagocytic properties. Systemic disease spread may be initiated by breaks in skin or mucosal barriers or by direct cellular invasion by GAS. Scarlet fever is a result of pyrogenic exotoxin production by the organism; pyrogenic exotoxins are also felt to play a principle role in triggering the deleterious immune activation of STSS.

Surface Virulence Factors

M protein is the single most studied and appreciated virulence feature of GAS. M proteins consist of two chains with a coiled-coil a-helical structure, very rare in bacteria, but a common motif in mammalian proteins. There are over 80 immunologically distinct M proteins on the basis of their hypervariable, negatively-charged N-termini; they also possess a conserved region, a proline/glycine rich region that intercalates the protein into the cell wall, and a hydrophobic membrane anchor region. Antigenic variation is due primarily to single amino acid substitutions. In general, the M protein types of GAS associated with impetigo and other skin infections are different from those associated with clinical pharyngitis. Rheumatic fever always follows pharyngitis and is more commonly associated with certain M types (1,3,5,16,18), although many serotypes have been implicated. Glomerulonephritis can follow pharyngitis (commonly M-types 1,4,12,15) or impetigo (M-types 49,52,55,59-61) and appears to be restricted to a smaller list of "nephritogenic" strains. Streptococcal toxic shock syndrome has been associated predominantly with M types 1 and 3. Induction of anti-M protein antibodies or cytotoxic T-cells which cross-react with cardiac myosin or basal ganglia neurons are felt to play a role in the pathogenesis of the carditis and chorea of rheumatic fever, while deposition of antigen-antibody complexes may play a role in the pathogenesis of acute poststreptococcal glomerulonephritis.

M proteins project from the cell wall as fimbriae and retard phagocytosis by host neutrophils and macrophages by inhibiting complement (C3b) deposition on the bacterial surface. The exact mechanism is somewhat controversial but probably involves binding of factor H to an exposed, conserved sequence in the C terminus of all M proteins. Factor H combines with C3b to make it susceptible to Factor I, which inactivates the alternative complement pathway C3 convertase. M protein may have other roles in pathogenesis, including directly or indirectly in mediating adherence to epithelial cells such as epidermal keratinocytes. M protein also binds fibrinogen. Fibrinogen can bind plasminogen that can be activated by bacterial streptokinase to form bound plasmin. Thus each bacterium could be coated itself with a host protease which may facilitate migration through host tissues.

The hyaluronic acid capsule of GAS is a polymer is essentially identical to human ground substance (? molecular mimicry to avoid immune detection). All pathogenic GAS have capsules, but some strains produce large capsules; these are often associated with more invasive infections and isogenic nonencapsulated mutant strains are less pathogenic in mice. The capsule may also mediate GAS adherence and paracellular tissue invasion via binding to the cellular hyaluronan receptor CD44.

F protein has been shown to be surface-expressed and to bind tightly to fibronectin, the extracellular matrix protein to which GAS bind. M protein is not expressed under conditions that promote expression of F protein (high O2 and low CO2), and vice versa. F protein may be the adhesin for respiratory epithelial cells but this is still controversial. F - mutants do not bind to Langerhans cells in the epidermis, but do bind normally to keratinocytes.

Lipotechoic acid (LTA) is found in abundance on the surface of GAS, adherent to M protein. There is some evidence that LTA is an adhesin that binds GAS to fibronectin, although this is controversial, and it may be that GAS has multiple adhesins that are used for attachment to different kinds of cells.

GAS also possess a family of immunoglobulin binding proteins which have structural similarities to M protein. These proteins can bind to the conserved Fc region of human IgG or IgA. This "non-immune" Ig binding prevents the immunoglobulin molecular from acting as an opsonin for neutrophils that bear Fc receptors on their surface.

C5a peptidase is a surface protein that inactivates C5a, a potent chemotactic peptide, generated by proteolytic activation of the fifth component of complement (C5). Its role in pathogenesis is unproven. Like the Ig-binding proteins, it is part of the same gene cluster (regulon) that encodes M protein.

The group A carbohydrate is anchored to the peptidoglycan in the cell wall; it stimulates antibodies, but they are not opsonic. This is the antigen that accounts for group specificity. It does not have a known role in virulence. The rapid group A antigen detection tests now in widespread clinical use for diagnosis of streptococcal pharyngitis are based on nitric acid extraction of the A polysaccharide from the bacterial cell wall and its detection with specific anti-group A antibody.

Secreted Virulence Factors

SpeA ribbon structure Streptococcal pyrogenic exotoxins (SPEs) A, B and C were formerly referred to as erythrogenic toxins. These toxins appear to be responsible for the rash of scarlet fever, and for the shock that may result from severe streptococcal infections. All three toxins induce fever, but are immunologically distinct. SPE B is a cysteine protease that is encoded on the chromosome of all GAS, but a few strains do not make an active protein. SPE B can cleave precursor interleukin-1B (pIL-1B) to its active form (pIL-1B is released by inflammatory macrophages), and can degrade fibronectin and vitronectin. SPE A and C are encoded by bacteriophage; their functions are not know but the return of severe GAS infection in the past decade was associated with the reappearance of SPE A producing bacteria. In one study 90% of GAS from patients with "serious disease" (shock of bloodstream invasion) made SPE A, whereas SPE A was found in only 54% of isolates from patients with uncomplicated infections. However, in a more recent survey SPE A was found in only 44% of isolates from patients with severe infections. The SPE proteins all have properties of superantigens, causing non-specific overactivation of host T-lymphocytes.

Streptolysin S (SLS) is an oxygen-stabile, non-antigenic, small molecular weight protein that may be primarily cell bound and is responsible for the ß-hemolytic phenotype produced by GAS on the surface of blood-agar plates. The gene encoding SLS has recently been identified and lies at the head of an operon of nine genes responsible for its activation and secretion to the cell surface. Streptolysin has structural similarities to other Gram-positive bacterial bacteriocins, a family of molecules which are toxic to other bacterial cell membranes and sometimes exhibit hemolytic or cytolytic properties. Thus SLS production may help GAS compete vs. other bacteria for a niche on epithelial surfaces. SLS appears necessary though not sufficient for GAS deep soft tissue infection, as SLS deficient mutants fail to produce necrotic skin lesions in the mouse model.

Streptolysin O (SLO) is an oxygen-labile molecular with weak hemolytic activity that polymerizes into rod-like structures that insert in cholesterol-containing membranes to form pores. SLO is cardiotoxic when infused intravenously into animals and SLO is antigenic. The antibody against SLO is often used to diagnose recent GAS infection

Streptokinase is a secreted protein that combines 1:1 with plasminogen to make functional plasmin, which can hydrolyze fibrin and other host proteins. When bound to the bacterial cell surface, plasmin is not inhibited by alpha-macroglobulin. In this way, streptokinase can promote spread of GAS through tissues. Streptokinase from Group G streptococcus is used therapeutically to dissolve clots. Streptokinase is antigenic and antibody can neutralize enzymatic activity.

Hyaluronidase dissolves ground substance and probably promotes movement of bacteria through tissue. The GAS capsule is also a substrate for this enzyme and it is not clear how the bacteria regulates synthesis and turnover of the two factors.

NADase and DNAases are made by most GAS. They have no known role in pathogenesis of infection but they are antigenic and can be used for serologic diagnosis.

Treatment

All GAS are exquisitely susceptible to penicillin. Most are also susceptible to erythromycin and clindamycin. For severe invasive GAS infections, many favor treatment with clindamycin, a ribosomally-active antibiotic which may decrease protein synthesis and elaboration of toxins. Intravenous gamma globulin has been used as an adjunctive therapy for invasive GAS infections with STSS.

Group B Streptococcus (S. agalactiae , GBS)

Para neustros amigos de habla español .... GBS should not be confused with ....

Phenotypic Appearance

GBS are ß-hemolytic but typically do not produce large zones of hemolysis such as those associated with streptolysin S of GAS. They also produce an orange pigment that can be used for clinical identification. Pigment production is linked to ß-hemolysin production, and both are accentuated by anaerobic incubation.

Epidemiology

GBS Bovine Mastitis GBS are found in cattle as well as humans, and they are an important etiologic agent of bovine mastitis. GBS commonly colonize the lower gastrointestinal tract and the vagina of healthy adults; the prevalence in the population at any given time is approximately 20-30%. GBS are the leading cause of pneumonia, sepsis and meningitis in newborn infants, who may acquire the infection from a colonized mother. The incidence of invasive neonatal GBS infection is approximately 1-3 cases per 1,000 live births, a number that is declining with more aggressive programs for screening pregnant women and administering intrapartum antibiotic prophylaxis. In recent years, GBS have been increasingly recognized as agents of invasive infections in adults, usually those who are immunocompromised, the elderly, parturient women, or diabet.

Clinical Manifestations

The more common early-onset form of GBS disease typically develops on the first day of life and is a fulminant infection involving pneumonia and septicemia. Premature infants are at especially high risk for severe early-onset GBS infection. The polysaccharide capsule serotypes associated with early-onset infection reflect their prevalence as colonizers in the adult population, with a relatively equal distribution among the common serotypes Ia, III, V and II. Late-onset GBS infection can occur in infants up to 6 months of age, often has a subacute presentation, and involves bacteremia and frequently meningitis. Type III GBS strains account for a disproportionately high percentage of late-onset cases and CNS infections.

GBS Clinical Images
Pathogenesis

GBS must first establish vaginal colonization in the pregnant mother by adhering to vaginal epithelial cells and resisting mucosal immune defenses. To gain access to the fetus, GBS may next ascend into the amniotic cavity by penetration of placental membranes. Chorioamnionitis and bacterial proliferation allow the bacteria to enter the fetal lung through aspiration of infected amniotic fluid. Alternatively, the infant may acquire the organism on passage through the birth canal. Pneumonia with lung epithelial and endothelial cell injury are characteristic of early onset disease, and may be mediated in part by the cytotoxic properties of ß-hemolysin and the influx of host neutrophils. GBS are capable of "invading" alveolar epithelial and pulmonary endothelial cells within membrane-bound vacuoles. This process may allow the organism to gain stepwise entry into the bloodstream. Host phagocytic defenses are subsequently called upon to clear the pathogenic bacteria. Newborn infants, and particularly premature infants, have fewer alveolar macrophages than adults and exhibit poor neutrophil chemotaxis. Furthermore, GBS are inefficiently phagocytosed in the absence of opsonization by specific antibody or complement, both of which may be present in diminished amounts in neonatal serum. The polysaccharide capsule of GBS has a marked inhibitory effect on phagocytic clearance by preventing complement deposition on the bacterial surface. Components of GBS surface protein C may both retard opsonization and decrease killing of GBS taken up by neutrophils. Cell wall-associated components of circulating GBS induce a sepsis syndrome characterized by severe systemic hypotension, pulmonary hypertension, hypoxemia and acidosis. This syndrome reflects the injurious effects of a host inflammatory response mediated by release of tumor necrosis factor (TNFalpha), interleukins, prostaglandins and thromboxane. Bloodstream dissemination allows GBS to reach multiple body sites, and invasion of brain microvascular endothelial cells may be the first step in production of meningitis.

Virulence Factors

GBS Capsule The GBS surface polysaccharide capsule is the most well-studied virulence determinant in neonatal infection, by virtue of its antiphagocytic properties. Capsule-deficient mutants have greatly diminished virulence in animal models. Sialic acid residues on the capsule inhibit the binding of opsonically-active C3 component of complement to the cell surface, thereby blocking activation of the alternative pathway. Transplacental passage of type-specific anticapsular IgG antibody from mother to infant is an important protective factor against invasive disease. Conjugate vaccines based on GBS polysaccharide capsules are protective for the newborn when administered to pregnant animals, with human trials now underway.

Specific mutant studies have shown that the GBS ß-hemolysin is cytotoxic to pulmonary epithelial and endothelial cells, and may contribute to the pulmonary injury and alveolar protein exudate of early-onset pneumonia. GBS hemolysin activity is blocked by surfactant phospholipid, providing a rationale for the increased risk of premature, surfactant-deficient newborns to severe pneumonia. This molecule also appears to induce cytokine release and nitric oxide production in macrophages and may stimulate elements of the sepsis cascade.

GBS C5a-peptidase specifically cleaves and inactivates the complement-derived neutrophil chemoattractant C5a. Animal model experiments show that C5a-peptidase-deficient mutants are more rapidly cleared from the lungs of infected animals when compared to the isogenic wild-type strain.

Treatment

Infants with GBS infection can be treated with penicillin, ampicillin or a cephalosporin. Universal screening of pregnant women at 35-37 weeks gestation for GBS colonization of the vagina or rectum, followed by prophylactic antibiotic therapy during labor to those women who test positive for GBS (plus those who have other risk factors such as fever, prolonged rupture of membranes, or premature delivery), has had a significant favorable impact on the incidence of early-onset GBS disease.

*Basic Microbiology* The genus Streptococcus is made up of non-motile, Gram-positive cocci that grow in chains because when they divide each cross wall is made perpendicular to the long axis of the developing chain. Metabolically, streptococci are anaerobes because they obtain energy by fermentation, even when grown in air. Some are facultative anaerobes while others are strict anaerobes. Because they are not able to synthesize cytochromes, they do not carry out oxidative phosphorylation. The principle end product of fermentation is lactic acid. Therefore, streptococci are more acid-tolerant than most bacteria, which affects their ecology. Another consequence of lacking cytochromes is the absence of the cytochrome-dependent enzyme catalase (2 H2O2 --> 2 H 2O + O2). This feature is useful for identifying streptococci since most other bacteria make catalase; it also explains why streptococci are so susceptible to killing by polymorphonuclear leukocytes that make hydrogen peroxide as a product of the respiratory burst. Nearly all medically important streptococci have multiple nutritional requirements (auxotrophies) that include vitamins, amino acids, and nucleic acids, and are therefore not free-living in the environment. This also means that streptococci grow only in nutritionally rich media, and colonies are usually small. *Classification of Streptococci* The most common classification system for streptococci used everyday in the clinical microbiology lab is based on the appearance of colonies grown on the surface of blood-agar media. As depicted in the photograph to the left, streptococci are designated as exhibiting either alpha, beta or gamma hemolytic activity. In so-called ß-hemolysis , the RBCs surrounding the colony are completely lysed. Specific protein "hemolysin" toxins are responsible for this phenotype. The important pathogens in this group are group A streptococci and group B streptococci. In the case of alpha-hemolysis , the RBCs are intact but their pigment is changed from red to green. Hydrogen peroxide production is responsible for this phenotype, and these streptococci are sometimes referred to as "viridans" (= green). The most important human pathogen in this group is Streptococcus pneumoniae . Gamma-hemolysis is in fact a misnomer, as there is no change in the appearance of the RBC. The ß-hemolytic streptococci are further classified on the basis of a scheme developed by Rebecca Lancefield (1895-1981) that tests the serologic reactivity of carbohydrate antigens (C substance) derived from acid extraction of the bacterial cell walls. Recognized serogroups are given letter designations from A-H and K-V. Some group antigens are shared by multiple species; however, only a single pathogenic species each comprises groups A (S. pyogenes) and B (S. agalactiae ). Other streptococci with pathogenic potential are found within the ß-hemolytic groups C, F and G (S. equisimilis or S. anginosus), but only extremely rarely is disease is associated with group D (S. bovis, S. durans or S. avium ) or other Lancefield groups. [Enterococcus faecalis and E. faecium were once classified as Group D streptococci]. S. pneumoniae (pneumococcus), a human pathogen producing pneumonia and meningitis, lacks a group specific antigen. Likewise, no group antigen is present in the viridans streptococci (e.g. S. mutans , S. sanguis, S. salivarius, S. milleri ) which are part of the oral flora and can contribute to the pathogenesis of dental caries or produce endocarditis on damaged heart valves. *Group A Streptococcus* ( S. pyogenes , GAS) Clinical Manifestations GAS cause infections of the upper respiratory tract (principally pharyngitis = "strep throat", cervical lymphadenitis, peritonsillar abscess, pneumonia and empyema), and the skin (impetigo, lymphangitis, cellulitis and erysipelas). In addition, GAS are the cause of scarlet fever, and two immunologically mediated diseases, rheumatic fever and acute glomerulonephritis. Since the mid-1980's, a significant increase in deep and systemic infections caused by GAS has been observed, in particular necrotizing fasciitis and septicemia, often complicated by development of streptococcal toxic shock syndrome (STSS) and multiorgan system failure. Watch a CNN News video clip describing invasive GAS infections including necrotizing fasciitis Copyright: (c) 2000 Cable News Network, an AOL Time Warner Company. All rights reserved (Requires the free RealPlayer software) Epidemiology GAS are exclusively human pathogens under natural conditions. Bacteria are passed from person to person, either by droplets, or by direct contact with infected skin, or possibly by fomites. Pharyngitis is more common in the winter and spring in temperate climates, and impetigo-like skin infections are more prevalent in the summer, often due to secondary infections of insect bites. The prevalence of GAS in the throats of school children varies from 5-40%. The highest incidence of GAS infections is in pre-adolescents. Skin trauma and chickenpox are important predisposing factors for severe, invasive GAS infections. Pathogenesis The pathogenesis of GAS infections begins with attachment to epithelial cells of the pharynx or skin via hairlike fimbriae that protrude from the cell surface and into the organism's hyaluronic acid capsule. The fimbriae are composed of lipotechoic acid and a series of serologically distinct surface proteins, the M proteins, which have important antiphagocytic properties. Systemic disease spread may be initiated by breaks in skin or mucosal barriers or by direct cellular invasion by GAS. Scarlet fever is a result of pyrogenic exotoxin production by the organism; pyrogenic exotoxins are also felt to play a principle role in triggering the deleterious immune activation of STSS. Surface Virulence Factors M protein is the single most studied and appreciated virulence feature of GAS. M proteins consist of two chains with a coiled-coil a-helical structure, very rare in bacteria, but a common motif in mammalian proteins. There are over 80 immunologically distinct M proteins on the basis of their hypervariable, negatively-charged N-termini; they also possess a conserved region, a proline/glycine rich region that intercalates the protein into the cell wall, and a hydrophobic membrane anchor region. Antigenic variation is due primarily to single amino acid substitutions. In general, the M protein types of GAS associated with impetigo and other skin infections are different from those associated with clinical pharyngitis. Rheumatic fever always follows pharyngitis and is more commonly associated with certain M types (1,3,5,16,18), although many serotypes have been implicated. Glomerulonephritis can follow pharyngitis (commonly M-types 1,4,12,15) or impetigo (M-types 49,52,55,59-61) and appears to be restricted to a smaller list of "nephritogenic" strains. Streptococcal toxic shock syndrome has been associated predominantly with M types 1 and 3. Induction of anti-M protein antibodies or cytotoxic T-cells which cross-react with cardiac myosin or basal ganglia neurons are felt to play a role in the pathogenesis of the carditis and chorea of rheumatic fever, while deposition of antigen-antibody complexes may play a role in the pathogenesis of acute poststreptococcal glomerulonephritis. M proteins project from the cell wall as fimbriae and retard phagocytosis by host neutrophils and macrophages by inhibiting complement (C3b) deposition on the bacterial surface. The exact mechanism is somewhat controversial but probably involves binding of factor H to an exposed, conserved sequence in the C terminus of all M proteins. Factor H combines with C3b to make it susceptible to Factor I, which inactivates the alternative complement pathway C3 convertase. M protein may have other roles in pathogenesis, including directly or indirectly in mediating adherence to epithelial cells such as epidermal keratinocytes. M protein also binds fibrinogen. Fibrinogen can bind plasminogen that can be activated by bacterial streptokinase to form bound plasmin. Thus each bacterium could be coated itself with a host protease which may facilitate migration through host tissues. The hyaluronic acid capsule of GAS is a polymer is essentially identical to human ground substance (? molecular mimicry to avoid immune detection). All pathogenic GAS have capsules, but some strains produce large capsules; these are often associated with more invasive infections and isogenic nonencapsulated mutant strains are less pathogenic in mice. The capsule may also mediate GAS adherence and paracellular tissue invasion via binding to the cellular hyaluronan receptor CD44. F protein has been shown to be surface-expressed and to bind tightly to fibronectin, the extracellular matrix protein to which GAS bind. M protein is not expressed under conditions that promote expression of F protein (high O2 and low CO2), and vice versa. F protein may be the adhesin for respiratory epithelial cells but this is still controversial. F - mutants do not bind to Langerhans cells in the epidermis, but do bind normally to keratinocytes. Lipotechoic acid (LTA) is found in abundance on the surface of GAS, adherent to M protein. There is some evidence that LTA is an adhesin that binds GAS to fibronectin, although this is controversial, and it may be that GAS has multiple adhesins that are used for attachment to different kinds of cells. GAS also possess a family of immunoglobulin binding proteins which have structural similarities to M protein. These proteins can bind to the conserved Fc region of human IgG or IgA. This "non-immune" Ig binding prevents the immunoglobulin molecular from acting as an opsonin for neutrophils that bear Fc receptors on their surface. C5a peptidase is a surface protein that inactivates C5a, a potent chemotactic peptide, generated by proteolytic activation of the fifth component of complement (C5). Its role in pathogenesis is unproven. Like the Ig-binding proteins, it is part of the same gene cluster (regulon) that encodes M protein. The group A carbohydrate is anchored to the peptidoglycan in the cell wall; it stimulates antibodies, but they are not opsonic. This is the antigen that accounts for group specificity. It does not have a known role in virulence. The rapid group A antigen detection tests now in widespread clinical use for diagnosis of streptococcal pharyngitis are based on nitric acid extraction of the A polysaccharide from the bacterial cell wall and its detection with specific anti-group A antibody. Secreted Virulence Factors Streptococcal pyrogenic exotoxins (SPEs) A, B and C were formerly referred to as erythrogenic toxins. These toxins appear to be responsible for the rash of scarlet fever, and for the shock that may result from severe streptococcal infections. All three toxins induce fever, but are immunologically distinct. SPE B is a cysteine protease that is encoded on the chromosome of all GAS, but a few strains do not make an active protein. SPE B can cleave precursor interleukin-1B (pIL-1B) to its active form (pIL-1B is released by inflammatory macrophages), and can degrade fibronectin and vitronectin. SPE A and C are encoded by bacteriophage; their functions are not know but the return of severe GAS infection in the past decade was associated with the reappearance of SPE A producing bacteria. In one study 90% of GAS from patients with "serious disease" (shock of bloodstream invasion) made SPE A, whereas SPE A was found in only 54% of isolates from patients with uncomplicated infections. However, in a more recent survey SPE A was found in only 44% of isolates from patients with severe infections. The SPE proteins all have properties of superantigens, causing non-specific overactivation of host T-lymphocytes. Streptolysin S (SLS) is an oxygen-stabile, non-antigenic, small molecular weight protein that may be primarily cell bound and is responsible for the ß-hemolytic phenotype produced by GAS on the surface of blood-agar plates. The gene encoding SLS has recently been identified and lies at the head of an operon of nine genes responsible for its activation and secretion to the cell surface. Streptolysin has structural similarities to other Gram-positive bacterial bacteriocins, a family of molecules which are toxic to other bacterial cell membranes and sometimes exhibit hemolytic or cytolytic properties. Thus SLS production may help GAS compete vs. other bacteria for a niche on epithelial surfaces. SLS appears necessary though not sufficient for GAS deep soft tissue infection, as SLS deficient mutants fail to produce necrotic skin lesions in the mouse model. Streptolysin O (SLO) is an oxygen-labile molecular with weak hemolytic activity that polymerizes into rod-like structures that insert in cholesterol-containing membranes to form pores. SLO is cardiotoxic when infused intravenously into animals and SLO is antigenic. The antibody against SLO is often used to diagnose recent GAS infection Streptokinase is a secreted protein that combines 1:1 with plasminogen to make functional plasmin, which can hydrolyze fibrin and other host proteins. When bound to the bacterial cell surface, plasmin is not inhibited by alpha-macroglobulin. In this way, streptokinase can promote spread of GAS through tissues. Streptokinase from Group G streptococcus is used therapeutically to dissolve clots. Streptokinase is antigenic and antibody can neutralize enzymatic activity. Hyaluronidase dissolves ground substance and probably promotes movement of bacteria through tissue. The GAS capsule is also a substrate for this enzyme and it is not clear how the bacteria regulates synthesis and turnover of the two factors. NADase and DNAases are made by most GAS. They have no known role in pathogenesis of infection but they are antigenic and can be used for serologic diagnosis. Treatment All GAS are exquisitely susceptible to penicillin. Most are also susceptible to erythromycin and clindamycin. For severe invasive GAS infections, many favor treatment with clindamycin, a ribosomally-active antibiotic which may decrease protein synthesis and elaboration of toxins. Intravenous gamma globulin has been used as an adjunctive therapy for invasive GAS infections with STSS. Group B Streptococcus (S. agalactiae , GBS) Para neustros amigos de habla español .... GBS should not be confused with .... Phenotypic Appearance GBS are ß-hemolytic but typically do not produce large zones of hemolysis such as those associated with streptolysin S of GAS. They also produce an orange pigment that can be used for clinical identification. Pigment production is linked to ß-hemolysin production, and both are accentuated by anaerobic incubation. Epidemiology GBS are found in cattle as well as humans, and they are an important etiologic agent of bovine mastitis. GBS commonly colonize the lower gastrointestinal tract and the vagina of healthy adults; the prevalence in the population at any given time is approximately 20-30%. GBS are the leading cause of pneumonia, sepsis and meningitis in newborn infants, who may acquire the infection from a colonized mother. The incidence of invasive neonatal GBS infection is approximately 1-3 cases per 1,000 live births, a number that is declining with more aggressive programs for screening pregnant women and administering intrapartum antibiotic prophylaxis. In recent years, GBS have been increasingly recognized as agents of invasive infections in adults, usually those who are immunocompromised, the elderly, parturient women, or diabet. Clinical Manifestations The more common early-onset form of GBS disease typically develops on the first day of life and is a fulminant infection involving pneumonia and septicemia. Premature infants are at especially high risk for severe early-onset GBS infection. The polysaccharide capsule serotypes associated with early-onset infection reflect their prevalence as colonizers in the adult population, with a relatively equal distribution among the common serotypes Ia, III, V and II. Late-onset GBS infection can occur in infants up to 6 months of age, often has a subacute presentation, and involves bacteremia and frequently meningitis. Type III GBS strains account for a disproportionately high percentage of late-onset cases and CNS infections. Pathogenesis GBS must first establish vaginal colonization in the pregnant mother by adhering to vaginal epithelial cells and resisting mucosal immune defenses. To gain access to the fetus, GBS may next ascend into the amniotic cavity by penetration of placental membranes. Chorioamnionitis and bacterial proliferation allow the bacteria to enter the fetal lung through aspiration of infected amniotic fluid. Alternatively, the infant may acquire the organism on passage through the birth canal. Pneumonia with lung epithelial and endothelial cell injury are characteristic of early onset disease, and may be mediated in part by the cytotoxic properties of ß-hemolysin and the influx of host neutrophils. GBS are capable of "invading" alveolar epithelial and pulmonary endothelial cells within membrane-bound vacuoles. This process may allow the organism to gain stepwise entry into the bloodstream. Host phagocytic defenses are subsequently called upon to clear the pathogenic bacteria. Newborn infants, and particularly premature infants, have fewer alveolar macrophages than adults and exhibit poor neutrophil chemotaxis. Furthermore, GBS are inefficiently phagocytosed in the absence of opsonization by specific antibody or complement, both of which may be present in diminished amounts in neonatal serum. The polysaccharide capsule of GBS has a marked inhibitory effect on phagocytic clearance by preventing complement deposition on the bacterial surface. Components of GBS surface protein C may both retard opsonization and decrease killing of GBS taken up by neutrophils. Cell wall-associated components of circulating GBS induce a sepsis syndrome characterized by severe systemic hypotension, pulmonary hypertension, hypoxemia and acidosis. This syndrome reflects the injurious effects of a host inflammatory response mediated by release of tumor necrosis factor (TNFalpha), interleukins, prostaglandins and thromboxane. Bloodstream dissemination allows GBS to reach multiple body sites, and invasion of brain microvascular endothelial cells may be the first step in production of meningitis. Virulence Factors The GBS surface polysaccharide capsule is the most well-studied virulence determinant in neonatal infection, by virtue of its antiphagocytic properties. Capsule-deficient mutants have greatly diminished virulence in animal models. Sialic acid residues on the capsule inhibit the binding of opsonically-active C3 component of complement to the cell surface, thereby blocking activation of the alternative pathway. Transplacental passage of type-specific anticapsular IgG antibody from mother to infant is an important protective factor against invasive disease. Conjugate vaccines based on GBS polysaccharide capsules are protective for the newborn when administered to pregnant animals, with human trials now underway. Specific mutant studies have shown that the GBS ß-hemolysin is cytotoxic to pulmonary epithelial and endothelial cells, and may contribute to the pulmonary injury and alveolar protein exudate of early-onset pneumonia. GBS hemolysin activity is blocked by surfactant phospholipid, providing a rationale for the increased risk of premature, surfactant-deficient newborns to severe pneumonia. This molecule also appears to induce cytokine release and nitric oxide production in macrophages and may stimulate elements of the sepsis cascade. GBS C5a-peptidase specifically cleaves and inactivates the complement-derived neutrophil chemoattractant C5a. Animal model experiments show that C5a-peptidase-deficient mutants are more rapidly cleared from the lungs of infected animals when compared to the isogenic wild-type strain. Treatment** Infants with GBS infection can be treated with penicillin, ampicillin or a cephalosporin. Universal screening of pregnant women at 35-37 weeks gestation for GBS colonization of the vagina or rectum, followed by prophylactic antibiotic therapy during labor to those women who test positive for GBS (plus those who have other risk factors such as fever, prolonged rupture of membranes, or premature delivery), has had a significant favorable impact on the incidence of early-onset GBS disease.

*Streptococci: The World's Favorite Bacteria*

Basic
Microbiology Classification of
Streptococci Group A
Streptococcus Group B
Streptococcus