STREPTOCOCCUS

&

ENTEROCOCCUS

REVIEW

Bacterial Cell Morphology• Gram Stain• Cytoplasmic (plasma) membrane • Cell wall structure• Bacterial cell shapes

Common Cell Membrane

Gram-Positive Cell Wall

Gram-Negative

Gram-Positive

Peptidoglycan

Teichoic Acid

Gram-Positive Cell Wall

Gram-Negative Cell Wall

Gram-Negative Cell Wall

Genus Streptococcus

Commensals or Parasites of man & animals or

Saprophytes of decaying matter

MorphologyGram-Positive Cocci in Pairs or Chains

Gram-PositiveStreptococcus

Genus Streptococcus

Physiology & MetabolismFacultative AnaerobesFastidious Growth RequirementsFermentative Metabolism of Carbohydrates:

Lactic acid, ethanol, acetate endproducts produced; No gasCatalase Negative (2H2O2 ---> O2 + 2H2O)

Separation of streptococci from staphylococciOxidase Negative (oxidoreductase oxidizes substrate w/ O2)

Beta, Alpha, or Gamma Hemolysis on blood agar

Genus StreptococcusRebecca Lancefield

Developed useful serogrouping system Classification of beta-hemolytic streptococci by group-specific cell wall carbohydrate (CHO) antigen As of 1992, Serogroups A to H and K to V

Groups A, B, C, D, and G are most comonly associated with human diseaseViridans streptococci and Streptococcus pneumoniae have no group-specific antigen

Antigenic StructureStreptococcus pyogenes (Group A)

Lancefield Group-specific antigen (C polysaccharide)Complex polysaccharide in cell wall

Proteins: Two major classes, M & T antigens Two minor classes, R & F

M-Protein: Type-specific antigen Fimbriae-like, hairy extensions

Resistant to heat and acid Trypsin Sensitive

Specific adherence by lipoteichoic acid and M-protein • (LTA-M) complexes

Antigenic StructureStreptococcus pyogenes (Group A)

Lancefield Group-specific antigen (C polysaccharide)Complex polysaccharide in cell wall

Proteins: Two major classes, M & T antigens Two minor classes, R & F

M-Protein: Type-specific antigen Fimbriae-like, hairy extensions

Resistant to heat and acid Trypsin Sensitive

Specific adherence by lipoteichoic acid and M-protein (LTA-M) complexes

T Antigens (not virulence factor)Resistant to trypsin, heat and acid; Adjunct to M-typing; Routine surveillance

Others

Antigenic Structure (cont.) Streptococcus pyogenes (Group A)

Capsular Polysaccharide: Hyaluronic acidNot present in all strainsSame as host hyaluronic acid (cartilage,skin etc)NonimmunogenicAntiphagocyticHyaluronidase (cell wall division) during late growth

Lipoteichoic Acid

Lancefield Serogroup Classification of Beta-Hemolytic Streptococci Important in

Human Disease

Group A Streptococci:Streptococcus pyogenes

• One of Most Important Human Pathogens• Suppurative Diseases:

Pharyngitis; Scarlet Fever; Cutaneous & Soft Tissue InfectionsSystemic Disease

• Non-Suppurative Sequelae:ARF,RHD,AG

Streptococcus pyogenes

(Phase Contrast)

Lancefield Serogroup Classification of Beta-Hemolytic Streptococci Important in

Human Disease (cont.)

Group B Streptococci: Streptococcus agalactiae

•Neonatal disease & obstetric complications

•Systemic, Cutaneous, UTI's

Streptococcus agalactiae

Lancefield Classification of Beta-Hemolytic Streptococci (cont.)

Group C Streptococci: Pharyngitis

Group G Streptococci:S.anginosus-milleri grp; Streptococcus spp.Pharyngitis Non-Lancefield Group Streptococci

Viridans StreptococciDental Caries: Streptococcus mutansStreptococcus sanguis; Streptococcus salivarius; Streptococcus mitis

Streptococcus pneumoniae

Enterococcus & Group D StreptococciGenitourinary Tract Infections (UTIs)Endocarditis

Major Human Diseases of Beta-Hemolytic Streptococci

Group A Streptococcus (S. pyogenes):Diverse group of acute suppurative (pus-forming) & nonsuppurative diseases

Suppurative Streptococcal DiseasesPharyngitis (& tonsilitis):

Scarlet fever: Complication of streptococcal pharyngitis when infecting strain is lysogenized; Frequently develop scarletina rash on upper chest spreading to extremities

Cutaneous & Soft Tissue Infxns.Pyoderma (Impetigo: contagious pyoderma with superficial yellow weeping lesions)Erysipelas: Acute superficial cellulitis of skin with lymphatic involvement; face and lower extremities, skin and subcutaneous tissues

Erysipelas

NOTE:√ erythema

√ bullae

Major Human Diseases of Beta-Hemolytic Streptococci (cont.)

Group A Streptococcus (S. pyogenes)Suppurative Streptococcal Diseases Cutaneous & Soft Tissue Infxns(cont.)

Cellulitis: Involvement of deeper subcutaneous tissues; Deeper invasion with systemic symptoms

Necrotizing fasciitis: (a.k.a., “flesh-eating bacteria”): Infection deep in subcutaneous tissues that spreads along fascial planes, destroying muscle and fat; Initially cellulitis followed by bullae (fluid filled blisters; bulla is singular), gangrene, systemic toxicity, multiorgan failure and mortality in more than 50% of patients

Wound Infections

Suppurative Streptococcal Diseases Group A Streptococcus (cont.)

Other Suppurative DiseasesPuerperal & neonatal sepsisLymphangitis: Inflammation of lymphatic vessel(s)Pneumonia

Systemic DiseaseStreptococcal Toxic Shock

Syndrome (TSS): Multisystem toxicity following soft tissue infection progressing to shock and organ failure (not to be confused with Staphylococcal Toxic Shock Syndrome where hyperabsorbent tampons have been identified as an important risk factor)

Bacteremia

Group A Streptococcal Diseases (cont.)

Nonsuppurative Sequelae Post-infection complications of Group A streptococcal disease; Serious complications in pre-antibiotic era; still important in developing countries Acute rheumatic fever (ARF):

Inflammation of heart, joints, blood vessels, sub-cutaneous tissues

Rheumatic heart disease (RHD): Chronic, progressive heart valve damage

Acute glomerulonephritis (AG): Acute inflammation of renal (kidney) glomeruli

Foodborne Disease

Epidemiology of Acute Streptococcal Infection• Predilection for upper respiratory tract or skin•Group A commonly colonize oropharynx of healthy children• M-types of strains colonizing throat differ from those on skin• Rapidly killed after phagocytosis, but cell walls not digested and may lead to chronic inflammatory lesions

Pharyngitis transmitted by droplets from respiratory secretions• Crowding increases risk (e.g., classrooms, day care facilities)

Pyoderma transmitted by direct contact with infectious lesions

Nonsuppurative Sequelae of Acute Group A Streptococcal Infection

Acute Rheumatic Fever (ARF)Inflammatory reaction characterized by arthritis, carditis, chorea (disorder of CNS with involuntary spastic movements), erythema marginatum (skin redness with defined margin), or subcutaneous nodulesWithin 2-3 weeks following pharyngitis• Epidemic pharyngitis: ARF in as many as 3%• Sporadic pharyngitis: ARF in 1 per 1000

Morbidity & mortality linked to subsequent disease of heart valve (Rheumatic Heart Disease)

Poorly understood pathogenesis with several proposed theories including cross-reactivity of heart tissues & strep AGNs •?? (Type ?? hypersensitivity, exotoxins, direct invasion)II

Nonsuppurative Sequelae of Acute Group A Streptococcal Infection (cont.)

Acute Glomerulonephritis Follows either respiratory (pharyngitis) or cutaneous (pyoderma) streptococcal infectionAssociated with well-defined group of M-types Incidence varies from <1% to 10-15%Most often seen in children manifesting as dark, smoky urine with RBC's, RBC casts, white blood cells, depressed serum complement, decreased glomerular filtration rateLatent period: 1-2 weeks after skin infection and 2-3 weeks after pharyngitisGranular accumulations of immunoglobulin due to deposition of immune complexes within the kidney (Type ?? Hypersensitivity)III

Determinants of PathogenicityCellular Virulence Factors

Capsule Antiphagocytic; Nonspecific adherenceHyaluronic acid (polysaccharide) mimics animal tissue

Lipoteichoic AcidCytotoxic for wide variety of cellsAdherence: Complexes with M protein (LTA-M) and binds to fibronectin on epithelial cells

M-ProteinLTA-M protein is adhesinAntiphagocyticInhibits alternate C’ pathway and opsonization

M-like Proteins: bind IgM and IgGF Protein: mediates adherence

Extracellular Virulence FactorsExotoxins:

Streptolysin O (SLO): Hemolytic and Cytolytic Prototype of oxygen-labile and thiol-activated cytolytic exotoxins (e.g., Streptococcus, Bacillus, Clostridium, Listeria)Lytic for variety of cells: bind to cholesterol-containing membranes and form arc- or ring- shaped oligomers that make cell leaky (RBC's, WBC’s, PMN's, platelets, etc.) Causes sub-surface hemolysis on BAPStimulate release of lysosomal enzymesSLO titer indicates recent infection (300-500 in pediatric populations)

Extracellellular Virulence Factors (cont.)

Exotoxins (cont.):Streptolysin S (SLS):

Hemolytic and CytolyticOxygen stable, non-antigenicLytic for red and white blood cells and wall-less forms (protoplast, L- forms)Causes surface hemolysis on BAP

Lysogeny: Lysogenized bacteriophages play key role in directing synthesis of various Group A streptococcal enzymes and toxins

• Pyrogenic Exotoxin (erythrogenic toxin)

• Phage-associated muralysins (lyse cell walls) produced by both Groups A and C

Extracellular Virulence Factors (cont.)

Exotoxins (cont):Pyrogenic (Erythrogenic) Exotoxins (Types A, B &C)

Produced by more than 90% of Grp A strep Lysogeny: Structural gene is carried by temperate bacteriophage, as is the case with diphtheria toxin

Mediate pyrogenicity (fever)Causes scarlet fever (scarletiniform) rashIncrease susceptibility to endotoxic shock

Type C toxin increases permeability of blood-brain barrierEnhance DTH Mitogenic for T lymphocytes (cause cell division), myocardial and hepatic necrosis, decrease in antibody synthesisImmunomodulators (superantigens): stimulate T cells to release cytokines

Cardiohepatic toxin

Extracellular Virulence Factors (cont.)

Enzymes:

Nucleases: Four antigenic types (A,B,C,D) Facilitate liquefication of pus generating growth substratesNucleases A, C have DNase activityNucleases B, D also have RNase activity

Streptokinases: Two different formsLyse blood clots: catalyze conversion of plasminogen to plasmin, leading to digestion of fibrin

C5a Peptidase: destroys C’ chemotactic signals (C5a)Hyaluronidase: hydrolyzes hyaluronic acidOthers: Proteinase, NADase, ATPase, phosphatase, etc.

Lab Identification of S. pyogenes (Group A)

• Primary culture by pour or streak plate• Domed,grayish/opalescent colonies

• Encapsulated cells produce mucoid colonies

Beta-hemolytic • Zone several times greater than diameter of colony

TSAS. pyogenes

Lab Identification of S. pyogenes (Group A) (cont.)

Catalase Negative: Differentiates from Staphylococcus

Bacitracin test: presumptively distinguishing between Group A beta-hemolytic streptococci (bacitracin POS) and other beta-hemolytic streptococci that are isolated from pharyngeal swabs (95% sensitivity for Grp A strep)

Rapid Identification Tests: Based on extraction of Group A carbohydrate directly from throat swabs

• ELISA, Coagglutination, Fluorescent Antibody

Group B Streptococcus

Streptococcus agalactiae

Group B Streptococcal Infections

Grp B Streptococcal Infections (cont.)

Age-Specific Attack Rates of Group B Streptococcal Disease

Epidemiology of Neonatal Group B Streptococcal Disease

Group B Streptococcus S. agalactiae

Diagnostic Laboratory Tests• CAMP factor positive

• Hippurase positive

CAMP Factor TestS. aureus

(Spingomyelinase C)Group B

Streptococcus(CAMP Factor)

Group A Streptococcus

Enhanced Zone of

Hemolysis

Hippurase NEG

Hippurase POS

Grp B Streptococci

and

Campylobacter

Streptococcus pneumoniae

• Commonly referred to as pneumococcus• Formerly Diplococcus pneumoniae

Streptococcus pneumoniae Infections

Epidemiology (cont.)Pneumococcal Infections (cont.)

S. pneumoniae• Diplococcus

S. pneumoniae: lancet-shaped diplococcus

S. pneumoniae Virulence Factors

S. pneumoniae Seasonal Incidence

Comparison of Morbidity & Mortality

for Bacterial Meningitis

Genetic Variation (Mutation)

Beginning of Molecular Genetics

Transformation (In vivo) (Griffith)

Streptococcus pneumoniae

Diagnostic Laboratory Tests• Optochin sensitivity (Taxo P disc)

Optochin SensitivityTaxo P

Streptococcus pneumoniae

Enterococcus faecalis Enterococcus faecium

• GI tract of humans and animals• Group D carbohydrate cell wall antigen• Formerly Streptococcus

Enterococcal Infections

Enterococcal Infections (cont.)

➢ Important nosocomial pathogen

➢ Vancomycin resistant Enterococcus (VRE)

Enterococcus

Diagnostic Laboratory Tests• Resistant to bile• Esculin hydrolysis• BEA media

Bile Esculin AgarPOS

EnterococcusGroup D Streptococcus

Bile Esculin AgarNEG

Esculin Bile

Assay

REVIEW

Lancefield Serogroup Classification of Beta-Hemolytic Streptococci Important in

Human DiseaseGroup A Streptococci:

Streptococcus pyogenesOne of Most Important Human PathogensSuppurative Diseases: Pharyngitis; Scarlet Fever;

Cutaneous & Soft Tissue Infections; Systemic Disease

Non-Suppurative Sequelae:ARF,RHD,AGGroup B Streptococci:

Streptococcus agalactiaeSystemic, Cutaneous, UTI's Neonatal diseaseObstetric Complications REVIEW

Nonsuppurative Sequelae of Acute Group A Streptococcal Infection

Acute Rheumatic Fever (ARF)Inflammatory reaction characterized by arthritis, carditis, chorea (disorder of CNS with involuntary spastic movements), erythema marginatum (skin redness with defined margin), or subcutaneous nodulesWithin 2-3 weeks following pharyngitis• Epidemic pharyngitis: ARF in as many as 3%• Sporadic pharyngitis: ARF in 1 per 1000

Morbidity & mortality linked to subsequent disease of heart valve (Rheumatic Heart Disease)

Poorly understood pathogenesis with several proposed theories including cross-reactivity of heart tissues & strep AGNs •?? (Type ?? hypersensitivity, exotoxins, direct invasion)II REVIEW

Nonsuppurative Sequelae of Acute Group A Streptococcal Infection (cont.)

Acute Glomerulonephritis Follows either respiratory (pharyngitis) or cutaneous (pyoderma) streptococcal infectionAssociated with well-defined group of M-types Incidence varies from <1% to 10-15%Most often seen in children manifesting as dark, smoky urine with RBC's, RBC casts, white blood cells, depressed serum complement, decreased glomerular filtration rateLatent period: 1-2 weeks after skin infection and 2-3 weeks after pharyngitisGranular accumulations of immunoglobulin due to deposition of immune complexes within the kidney (Type ?? Hypersensitivity)III REVIEW

Determinants of PathogenicityCellular Virulence Factors

Capsule Antiphagocytic; Nonspecific adherenceHyaluronic acid (polysaccharide) mimics animal tissue

Lipoteichoic AcidCytotoxic for wide variety of cellsAdherence: Complexes with M protein (LTA-M) and binds to fibronectin on epithelial cells

M-ProteinLTA-M protein is adhesinAntiphagocyticInhibits alternate C’ pathway and opsonization

M-like Proteins: bind IgM and IgGF Protein: mediates adherence REVIEW

Extracellular Virulence FactorsExotoxins:

Streptolysin O (SLO): Hemolytic and Cytolytic Prototype of oxygen-labile and thiol-activated cytolytic exotoxins (e.g., Streptococcus, Bacillus, Clostridium, Listeria)Lytic for variety of cells: bind to cholesterol-containing membranes and form arc- or ring- shaped oligomers that make cell leaky (RBC's, WBC’s, PMN's, platelets, etc.) Causes sub-surface hemolysis on BAPStimulate release of lysosomal enzymesSLO titer indicates recent infection (300-500 in pediatric populations) REVIEW

Extracellellular Virulence Factors (cont.)

Exotoxins (cont.):

Streptolysin S (SLS): Hemolytic and CytolyticOxygen stable, non-antigenicLytic for red and white blood cells and wall-less forms (protoplast, L- forms)Causes surface hemolysis on BAP

REVIEW

Extracellular Virulence Factors (cont.)

Exotoxins (cont):Pyrogenic (Erythrogenic) Exotoxins (Types A, B &C)

Produced by more than 90% of Grp A strep Lysogeny: Structural gene is carried by temperate bacteriophage, as is the case with diphtheria toxin

Mediate pyrogenicity (fever)Causes scarlet fever (scarletiniform) rashIncrease susceptibility to endotoxic shock

Type C toxin increases permeability of blood-brain barrierEnhance DTH Mitogenic for T lymphocytes (cause cell division), myocardial and hepatic necrosis, decrease in antibody synthesisImmunomodulators (superantigens): stimulate T cells to release cytokines

Cardiohepatic toxin REVIEW

Extracellular Virulence Factors (cont.)

Enzymes:

Nucleases: Four antigenic types (A,B,C,D) Facilitate liquefication of pus generating growth substratesNucleases A, C have DNase activityNucleases B, D also have RNase activity

Streptokinases: Two different formsLyse blood clots: catalyze conversion of plasminogen to plasmin, leading to digestion of fibrin

C5a Peptidase: destroys C’ chemotactic signals (C5a)Hyaluronidase: hydrolyzes hyaluronic acidOthers: Proteinase, NADase, ATPase, phosphatase, etc.

REVIEW

Epidemiology of Neonatal Group B Streptococcal Disease

REVIEW

REVIEW

Streptococcus pneumoniae Infections

REVIEW

➢Infections from endogenous spread from naso- or orapharynx

➢Pneumonia; sinusitis; otitis media; bacteremia; meningitis

➢Colonization highest in children

➢Antecedent viral respiratory tract disease increases risk

➢Most common in cold months

➢Polyvalent vaccine available (newly available for children)

S.pneumoniae Virulence Factors

REVIEW

Comparison of Morbidity & Mortality

for Bacterial Meningitis

REVIEW

Genetic Variation (Mutation)

REVIEW

Beginning of Molecular Genetics

REVIEW

Transformation (In vivo) (Griffith)

REVIEW

Enterococcal Infections

REVIEW

➢Group D cell wall antigen

➢Enterococcus faecalis; Enterococcus faecium

➢GI tract of humans and animals

➢UTI most common; wound infections; bacteremia; endocarditis

➢Most infections from endogenous source

➢Prolonged hospitalization and broad-spectrum antibiotics increase risk

➢Antibiotic resistance (VRE)