Streptococcus and Enterococcus

• Streptococcus and Enterococcus

  Family: Streptococcaceae
• Streptococcus

  • Gram (+) cocci in pairs in chains
  • A weak false (+) catalase in media with blood (peroxidase activity of hemoglobin)
  • Appear more elongated in chains when growth in broth cultures
  • Facultative anaerobes, aerotolerant anaerobes, capnophilic
  • Can ferment glucose (lactic acid), no gas
  • Colonies: small and transparent
• Lancefield Classification

  Rebecca Lancefield (1930) C carbohydrate (polysaccharide) in the cell wall
• Streptococcus pyogenes (Group A Streptococcus) has 8 virulence factors
• Virulence Factors of Streptococcus pyogenes

  • M protein (encoded by emm genes, resist phagocytosis & adhere to mucosal cells, 80 serotypes)
  • Protein F (fibronectin-binding protein) & Lipoteichoic acid (adherence to epithelial cells)
  • Hyaluronic acid capsule (prevents opsonized phagocytosis, masks antigens)
  • Streptolysin O (anaerobic hemolysis) and Streptolysin S (aerobic hemolysis, lyse leukocytes, platelets, RBCs)
  • Deoxyribonuclease (DNase A, B, C, D, most common: DNase B, immunogenic)
  • Streptokinase (lysis of fibrin clots through plasminogen activation, immunogenic)
  • Hyaluronidase (spreading factor, solubilizes ground substance)
  • Streptococcal Pyrogenic Exotoxins (cause scarlet fever)
• Infections caused by Streptococcus pyogenes

  • Bacterial pharyngitis
  • Pyodermal infections (impetigo, cellulitis, erysipelas, wound infection, arthritis, scarlet fever)
  • Necrotizing Fasciitis (Type 2)
  • Streptococcal Toxic Shock Syndrome
  • Poststreptococcal sequelae (rheumatic fever, acute glomerulonephritis)
• Streptococcus agalactiae (Group B Streptococcus)

  Virulence factors: Capsule (resist phagocytosis), Hemolysin, CAMP factor, neuraminidase, DNase, hyaluronidase, protease
• Biochemical Tests

  1. Catalase (no or few bubble formation for streptococci and enterococci)

  2. Bacitracin test (Taxo A) (zone of inhibition for S. pyogenes)

  3. Pyrrolidonyl-α-Nathylamide Hydrolysis Test (PYR Test) (bright red color for S. pyogenes)

  4. Hippurate Hydrolysis Test (deep purple color for S. agalactiae)

  5. CAMP Test (enhanced hemolysis for S. agalactiae)
• Streptococcus pneumoniae

  pneumococcus

  Member of S. mitis group,

  cell wall: C substance reacts with CRP to form precipitate
• Virulence Factors of Streptococcus pneumoniae

  • Capsular polysaccharide (susceptible to opsonization, Strains that lack capsule: non-pathogenic)
  • hemolysin, immunoglobulin A protease, neuraminidase and hyaluronidase
  • TOC: Penicillin
• Streptococcus pneumoniae is the #1 cause of pneumonia in ICP patients, also causes sinusitis, otitis media, bacteremia, and meningitis

  most frequently encountered in children < 3 years old with recurrent otitis media
• Pneumococcal Vaccines

  • PCV7 (7 serotypes conjugated to diphtheria protein, for children)
  • PCV13 (additional 6 serotypes, for children <5 years old)
  • PS23 (23 purified capsular polysaccharides)
• Specimen collection for Streptococcus pneumoniae

  RT specimens
• Direct microscopic examination of Streptococcus pneumoniae

  1. Effusions: Gram (+) pneumococci with numerous WBC

  2. CSF: Gram (+) cocci in pairs with numerous WBC
• Culture of Streptococcus pneumoniae

  1. BHI, TSB with 5% sheep RBCs or CAP

  2. 18 to 24 hours of incubation at 35° C to 37° C with increased CO2 for growth during primary isolation
• Streptococcus pneumoniae colonies

  • Round, glistening, wet, mucoid, dome-shaped appearance with large zone of α-hemolysis on SBA surrounding the colonies

  
• Streptococcus pneumoniae microscopic examination

  • Gram (+) cocci in pairs or in singly or in short chains
  • Ends of the cells are slightly pointed, giving them an oval or lancet shape

  
• Optochin test for Streptococcus pneumoniae

  1. Principle: determine the effect of optochin on an organism

  2. Result: (+) Zone of inhibition is 14 mm or greater in diameter, with 6-mm disk

  3. Result: (-) No zone of inhibition other α-hemolytic streptococcus
• Bile solubility test for Streptococcus pneumoniae

  1. Principle: bile or a solution of a bile salt rapidly lyses pneumococcal colonies

  Bile salts lowers the surface tension between the bacterial cell membrane and the medium.

  Accelerates the organism’s natural autolytic process intracellular

  autolytic enzyme.

  2. Result: (+) Lysed colonies (S. pneumoniae)

  3. Result: (-) Intact colonies (other α-hemolytic streptococcus)
• Viridans streptococci

  • Normal microbiota of the upper respiratory tract, the female genital tract, and the gastrointestinal tract
  • Viridans means "green" (α-hemolysis)
  • β-hemolytic and nonhemolytic species
  • Fastidious, with some strains requiring CO2 for growth
• Viridans streptococcus groups

  • S. mitis group
  • S. mutans group
  • S. salivarius group
  • S. bovis group (Group D antigen)
  • S. anginosus group (Lancefield group A, C, F, G, or N antigen)

  
• Viridans streptococci are the most common cause of subacute bacterial endocarditis
• Viridans streptococci cause oral infections such as gingivitis and dental caries
• Virulence factors of S. anginosus group

  • Polysaccharide capsule and cytolysin
  • Adherence and colonization in endocarditis
• Virulence factors of Groups C and G streptococci

  • M proteins, SLO, hyaluronidase, and DNase
• Viridans streptococcus colonies

  • Small and are surrounded by a zone of α-hemolysis
  • β-hemolytic or nonhemolytic

  
• Viridans streptococcus microscopic examination

  • Gram (+) cocci in pairs and in chains

  
• LAP test for viridans streptococci

  1. Principle: detection of the enzyme leucine aminopeptidase

  2. Substrate: Leucine-β-naphthylamide Hydrolyzed β-naphthylamine + paradimethyl aminocinnamaldehyde reagent

  3. Result: (+) Red color (Viridans streptococci)

  4. Result: (-) No color change or an slight yellow color (other α-hemolytic streptococcus)
• Enterococcus

  • Previously classified as group D streptococci
  • D antigen
  • Normal flora of intestinal tract of human and animals
  • Nonhemolytic, α-hemolytic or β-hemolytic
  • Ability to grow under extreme conditions (6.5% NaCl 45° C alkaline pH)
  • Pseudocatalase reaction
  • E. faecalis & E. faecium
  • E. durans, E. avium, E. casseliflavus, E. gallinarum, and E. raffinosus
  • PYR (+)

  
• Enterococcus faecalis virulence factors

  • Extracellular surface adhesin proteins, extracellular serine protease, and gelatinase (colonization and adherence)
  • Cytolysin (similar to bacteriocins produced by gram-positive bacteria)

  
• Enterococcus faecalis infections

  • Nosocomial infections (UTI & bacteremia)
  • Endocarditis in elderly patients
  • Intraabdominal or pelvic wound infections
• Specimen collection for Enterococcus

  Blood, urine, or wound specimens

  
• Culture of Enterococcus

  1. TSB or BHI with 5% sheep's blood

  2. Bile esculin azide, colistin– nalidixic acid, phenylethyl alcohol, chromogenic substrates, or cephalexin-aztreonam-arabinose agar

  3. 35° C in the presence of CO2 but do not require a high level of CO2 for growth

  
• Biochemical tests for Enterococcus

  1. Acid production in carbohydrate broth

  2. Arginine hydrolysis

  3. 0.04% tellurite tolerance

  4. Pyruvate utilization

  5. Resistance to 100-µg efrotomycin acid

  6. Positive motility

  
• Bile esculin test for Enterococcus

  1. Principle: Two-step test detecting growth of bacteria in the presence of 40% bile and the ability to hydrolyze esculin

  2. Result: (+) Blackening of the medium (Enterococcus spp. and Group D Streptococci)

  3. Result: (-) No color change (Viridans streptococcus)

  
• Salt tolerance test or 6.5% NaCl test for Enterococcus


  1. Principle: Ability to tolerate high salt environment

  2. Result: (+) Growth (turbidity) (Enterococcus and Pediococcus)

  3. Result: (-) No growth (clear) (Group D streptococci)
• Abiotrophia and Granulicatella spp.

  • Previously known as nutritionally variant streptococci
  • Require sulfhydryl compounds for growth
  • Oral and gastrointestinal microbiota
  • Bacteremia, endocarditis & otitis media
  • Satellitism on SBA with S.aureus
  • Media supplemented with 10 mg/L pyridoxal hydrochloride
• Biochemical tests for Abiotrophia and Granulicatella

  1. Production of: α-galactosidase, β-galactosidase, β-glucuronidase

  2. Hippurate hydrolysis

  3. Arginine hydrolysis

  4. Acid production from trehalose & starch
• Aerococcus

  • Common airborne organism
  • Opportunistic pathogen associated with bacteremia, endocarditis, and UTI in ICP
  • Similar to streptococci in culture and staphylococci in microscopic exam
  • Weak catalase or pseudocatalase
  • Growth in 6.5% NaCl
  • A. viridans: bile esculin & PYR (+)
  • A. urinae: bile esculin & PYR (-)
• Gemella

  • Similar to viridans streptococci in colonial morphology
  • α-hemolysis or nonhemolytic
  • Gram (-) cocci in pairs, tetrads, clusters, or short chains
  • Endocarditis, wounds, and abscesses
  • G. haemolysans