Streptococcus, Entrococcus and Other Catalayse Negative Gram Positive BacteriaAgustin, Aquitania, Bagsican

3E – Medical Technology

Microbiology

General Characteristics Streptococcus and Enterococcus

family Streptococcaceae

CATALAYSE GRAM NEGATIVE

GRAM POSITIVE COCCI – peptidoglycan & techoic acid

VIRIDANS GROUP layer: common C carbohydrate (polysaccharide) [+ antigen]

Usually pairs/chains (Broth)

Faculatative anaerobes

AEROTOLERANT ANAEROBES Grow in the presence of O2; unable in respiration

Production of Lactic Acid (Major End Product), fermentation

CAPNOPHILIC: ↑conc. of CO2

Media:

✔ ENRICHED with BLOOD & SERUM [small & transparent]

✖ POOR in NUTIENT MEDIA, TSA (Trypticase Soy Agar)

Classification Schemes

CATALAYSE GRAM NEGATIVE, GRAM POSITIVE COCCI

HEMOLYTIC PATTERN ON SBA

PHYSIOLOGIC CHARACTERISTICS

LANCEFIELD CLASSIFICATION

CHEMICAL CHARACTERISTICS

Hemolytic Patterns

Mahon, Diagnostic Microbiology 3rd. Ed., pg384

Hemolytic Patterns

•Partial lysis of RBC around colony•Greenish discolorationα•Complete lysis of area around colony•Clear area around colonyβ•No lysis of RBC•No change in agarNonhemolytic

•Small area of intact RBC around colony surrounded by a wider zone to complete hemolysis

Alpha-prime / Wide zone

Hemolytic Patterns

•Partial lysis of RBC around colony•Greenish discolorationα•Complete lysis of area around colony•Clear area around colonyβ•No lysis of RBC•No change in agarNonhemolytic

•Small area of intact RBC around colony surrounded by a wider zone to complete hemolysis

Alpha-prime / Wide zone

Physiologic Characteristics PYOGENIC STREPTOCOCCI

Pus-producing

Mostly beta-hemolytic

Majority of Lancefield groups

LACTOCOCCI Nonhemolytic with Lancefield group N

antigen

Dairy products

ENTEROCOCCI Normal flora of human intestine

VIRIDANS STREPTOCOCCI Widely found as normal flora in upper

respiratory tract in human

Lack C carbohydrate

Some have A, C, F, G, or N antigen

Alpha-hemolytic or Nonhemolytic

Opportunistic pathogens

Lancefield Classification Scheme Rebecca Lancefield (1930s)

Based on extraction of C carbohydrate from the streptococcal cell wall by placing the organism in dilute acid and heating for 10mins

Soluble antigen: immunize rabbits to obtain antisera to various C carbohydrate groups

Serologic groups designated by letters

DNA relatedness

Lancefield Classification SchemeClassification

Group A - Streptococcus pyogenes

Group B - Streptococcus agalactiae

Group C - Streptococcus equisimilis, Streptococcus equi, Streptococcus zooepidemicus, Streptococcus dysgalactiae

Group D - Enterococci, Streptococcus bovis

Group E - Streptococcus milleri and mutans

Group F - Streptococcus anginosus

Group G - Streptococcus canis, Streptococcus dysgalactiae

Group H - Streptococcus sanguis

Group L - Streptococcus dysgalactiae

Group N - Lactococcus lactis

Group R&S - Streptococcus suis

other Streptococcus species are classified as non-Lancefield Streptococci

Lancefield Classification SchemeSpecies

Lancefield Group

AntigenHemolysis

Type(s) Common Terms Disease Association(s)

S. pyogenes A β* Group A strep Rheumatic fever, scarlet fever, phayngitis, glomeruloneohritis, pyogenic infections

S. agalactiae B β* Group B strep Neonatal sepsis, meningitis, puerperal fever, pyogenic infections

S. equisimillis, S. equi subsp. Zooepidemicus, S. equi

C β* Group C strep Pharyngitis, urinal tract infections, pyogenic infections

S. bovis, S. equinus D α, none Nonenterococcus Endocartitis, urinary tract infections, pyogenic infections

E. faecalis, E. faecium, E. durans

D α, β, none

Enterococcus urinary tract infections, pyogenic infections

Other species F, G β (α, none)

Pheunococcus Pyogenic infections

S. pneumoniae - Pneumonia, meningitis, pyogenic infections

Anginosus group, Mutans group, Mitis group, Salivarius group

- α, none Viridans strep Emdocarditis, dental caries, abscesses in various tissues

Mahon, Diagnostic Microbiology 3rd. Ed., pg386

Biochemical Classification Can be performed in small lab

Definitive identification requires large number of biochemical characteristics or serological mtds., presumptive identification can be accomplished relatively easily with a few key tests & characteristics

Multitest commercial kits [Remel’s IDS Rapid STR]

Selecting identification scheme / kits Population served

Cost of an expanded identification scheme

Resources & abilities of the lab

Biochemical Classification Bacitracin Susceptibility

CAMP Test

Hippurate Hydrolysis

PYR Hydrolysis

Leucine Aminopeptidase

Vogues-Proskauer Test

Beta-D-Glucuronidase

Bile Esculin and Salt Tolerance

Optochin Susceptibility and Bile Solubility

Biochemical Classification Bacitracin Susceptibility

Identification of group A streptococcus

Sensitivity to bacitracin (pure culture only) >> S. pyogenes

Media: 5% SBA

Reagent: Bacitricin disk (0.04 units)

Incubation: 35°C overnight

Trimethprim & Sulfamethoxazole (SXT)

✔ [Suseptible] Any zone of inhibition

✖ [Resistant] Uniform lawn of groth to the edge of the disk

Mahon, Diagnostic Microbiology 3rd. Ed., pg390

Sucspetiblity of group A β-hemolytic strep to SXT

Biochemical Classification CAMP Test (Christie, Atkins and Munch Petersen)

3 Ways: Identification of group B streptococcus [enhanced hemolysis]

>> S. agalactiae

Β-lysin-producing strain of Staphylococcus aureus (arrowhead-shaped hemolysis)

Use of disk impregnated with β-lysin (arrowhead-shape hemolysis)

CAMP test

Media: 5% SBA

Incubation : 35°C @ least 2mins.

Controls: (+) S. agalactiae (-) S. pyogenes

✔ [Suseptible] Arrowhead-shaped area

✖ [Resistant] No enhanced hemolysis

→ CAMP inhibition rxn: Inhibition of hemolysis by S. aureus where 2 streaks approach each other [Arcanobacterium haemolyticum]

Biochemical Classification Hippurate Hydrolysis