STREPTOCOCCUS

CLASSIFICATION : By two ways:

1-Haemolytic Activity Classification: a)Beta-haemolytic streptococci:

complete haemolysis, decolorization of Hb, haemolysis more marked on anaerobiasis .

b) Alpha-haemolytic streptococci: partial haemolysis, a green brown colour due to reduction of Hb.

c) Non-haemolytic streptococci: no alpha or beta-haemolysis, Hb slightly discolourized.

2- Lancefield Classification: @ Beta haemolytic streptococci produce group specific Ags. C-carbohydrates.

@ Identified by group specific antiserum (Lancefield groping is A-H & K-V)

@ Most important beta-haemolytic S. pyogenes gp. A .

@ Other pathogenic streptococci belong to groups B, C, D, F, G.

STREPTOCOCCI

SPECIES & GROUPS:

Main species and groups are:

1. S. pyogenes (Lancefield gp. A).

2. S. agalactiae (Lancefield gp. B).

3. E. faecalis (Lancefield gp. D)

Other streptococci include:

4. S. viridans e.g.: S. mitis, S. mutans, S. sanguis5. Anaerobic streptococci.6. S. pneumoniae.

NORMAL HABITAT:@ Found in water, dust, milk, vegetation, milk products.

@ Commensal in URT: S. pyogenes (groups C, & G), & viridans strep.

@ Commensal in GIT: E. faecalis

@ Commensal female genitalia : Group B streptococci

PATHOGENICITY:

1- S. pyogenes (group A):

a) Tonsillitis & pharyngitis.

b) Peritonsilar abscess (quinsy).

c) Scarlet fever, caused by erythrogenic toxin in 2% of S. pyogenes strains.

d) Otitis media and mastoiditis.

e) Puerperal sepsis.

f) Skin infections, e.g.: cellulitis, erysipelas of face & scalp-

حمرة impetigo (blistering of skin).

g) Septicaemia, & endocarditis.

2- Post-streptococcal diseases:

Complications of acute group (A) streptococcal infection following an immunological response:

a)Acute glomerulonephritis, follows strep. skin infection .

b) Rheumatic fever, follows a respiratory strep.

infection, damaging of heart valves &

muscles.

3- Enzymes of S. pyogenes: a) Streptokinase: lyses fibrin .

b) Hyaluronidase, breaks down hyaluronic acid & makes S. pyogenes spread in tissues.

c) DNAses (A, B, C, D), break DNA.

d) DPNase, kills Leucocytes by attacking diphospho-pyridine nucleotide (DPN) in cell.

Toxins of S. pyogenes :

a)Streptolysin (O) : lyses RBC & stimulates production of

ASO .

b) Streptolysin (S) : lyses RBC

to give beta- haemolysis on B.A.

c) Erythrogenic toxin : causes skin

rash of scarlet fever.

4-S.agalactiae (group B)

Causes the following:

a)Neonatal septicaemia, pneumonia, meningitis.

b) Septic abortion, puerperal sepsis, vaginitis.

5- E. faecalis (group D)

Causes:

a) UTI, in association with

E. coli.

b) Ulcers, wound infections

c) Endocarditis, meningitis.

6- S. viridans: Causes the following:

a) Infective endocarditis, (in patients with damaged

heart valves).b) Dental caries.c) Abdominal abscess, Brain abscess.

7-S. pyogenes (groups C,G)

Causes the following:

a)Sinusitis bacteraemia.

b) Endocarditis urogenital infections.

c) Wound infections.

LABORATORY DIAGNOSIS

SPECIEMENS:1- Throat swabs, pus , blood (culture & ASO)

2- For S. agalactiae : HVS, blood, CSF, ear swabs (infants)

3- For E. faecalis: urine and pus

4- For streptococcal endocarditis: blood, urine

MICROSCOPY:

@ Streptococci : gram positive, non-motile, some strains are capsulated.

@ Morphology differs:

a) In fluid media: Long chains.b) In pus & solid media: Short chains, in pairs or single.

CULTURE:@ Grow aerobically & anaerobicaly. Temp. 15 - 45˚C .

@ Colonies on B.A. less than 1 mm. Colourless or grey - white dry, shiny, irregular outline.

@ Pathogenic streptococci produce haemolysis on B.A.

1- S. pyogenes (group A):

@ Shows beta-haemolysis on B.A. @ Sensitive to bacitracin

@ Other strep. species are also sensitive. @ Hence confirm by Lancefield grouping

@ Crystal violet B.A. selective for S. pyogenes & inhibits S. aureus.

@ Best medium to isolate S.pyogenes from throat & skin .

@ S. pyogenes will not grow on MacConkey

2- S. agalactiae (group B)

@ ß-hemolytic on B.A. @ Kanamycin B.A. is selective to isolate from urogenital sites @ On serum starch agar, It produces an orange pigment. @ Some strains grow on Mc Conkey agar.

CAMP Test:

@ S. agalactiae produces the protein CAMP (Christie, Atkins, & Munch Peterson) factor.

@ This factor interacts with staph. beta-haemolysin on RBC.

CAMP is performed as follows:

a) Streak Staph. across B.A plate, inoculate test organism at right angle without touching the

staph. b) Inoculate E. faecalis across

plate as a negative control.c) Incubate plate at 37˚C

overnight, look for interaction of test organism with staph.

Positive: identifies S. agalactiae.

CAMP Toxin method:

@ Add 2 drops of Staph. (B) toxin to a B.A. plate culture of S. agalactiae.

@ After 2 hrs incubation, development of haemolytic area around colonies indicates presence of S. agalactiae.

Test organism

Staph. Inoculum

Arrow-head of haemolysis produced by S.agalactiae

E. Faecalis Negative control

3- Beta-haemolytic group (C) & group (G)

streptococci.

@ Isolated from throat swabs and blood.

@ Identification is made by Lancefield grouping.

4- S. viridans :

@ Show α-haemolysis,

@ Optochin resistant

@ Not soluble in bile

5- E. faecalis (group D) :

@ Beta-haemolytic, alpha-haemolytic, or non-haemolytic.

@ Identified by positive rapid litmus milk reduction test.

@ On Mc Conkey, give small, dark red colonies.

Lancefield grouping

1- Earlier , grouping was done by a precipitation test applying heat & HCl to extract antigen of cell wall.

2- Test is latex, made by adding organism to antiserum containing latex particles on a slide .

@ Follow manufacturer procedure

BILE SOLUBILITY TEST:@ S.pneumoniae (soluble)-S. viridans (insoluble).

1- Emulsify colonies of test organism in a tube with 2 ml. distilled water.

2- Divide the suspension into two tubes.

3- To tube 1 add 2 drops sod. Deoxycholate, mix.

4- To tube 2 add 2 drops distilled water, mix.

5- Leave tubes for 15 min., look for clearing of turbidity in tube 1 .

Result: Clearing : soluble (Pneumococcus)

No clearing: insoluble (S. viridans).

LITUMS MILK DECOLORIZATION TEST:

@ E.faecalis decolorizes litmus milk.

1- Inoculate heavily test organism in 0.5 ml. litmus milk medium .

2- Incubate at 37˚C for 4 hrs., examine every ½ hr for reduction (litmus changes from purple to white or pale yellow).

3- Incubation not more than 4 hrs, since S. viridans reduces litmus after 4 hrs .

Result: Positive means E. faecalis.

ASO TITRE

@ To investigate post-streptococcal complications following Strep. infection

Other conditions giving rise in titre are:

Pneumococcal pneumonia, tuberculosis, gonorrhea, hepatitis, and rheumatoid

arthritis,group (C) & group (G) strep. infections

@ ASO is estimated by :1- Latex slide agglutination.2- Microtitration or tube haemolysis test.

DNAse ANTIBODY TEST:

@ To diagnose acute glomerulonephritis.

@ Use commercial kits.

STREPTOZYME TEST:

@ Available commercially.

@ A slide haemagglutination test to detect: * ASO, * Anti-DNAse B, * Anti-hyaluronidase, * Anti-NADase, * Anti-streptokinase.

ANTIMICROBIAL SENSITIVITY:

@ S.pyogenes & S. agalactiae: sensitive to: * penicillin * erythromycin.

@ S.pyogenes is resistant to: *polymyxin, *nalidixic acid, *sulphonamides.

S.PNEUMONIAE

SPECIES:

Only S.pneumoniae (pneumococcus)

NORMAL HABITAT: Commensal of URT .

PATHOGENICITY:

1- Lobar pneumonia, bronchitis.

bronchopneumonia,

2- Bacteraemia, meningitis.

3- Endocarditis, pericarditis.

4- Otitis media , sinusitis, conjunctivitis.

ENZYMES:

Hyaluronidase.

TOXINS:

Leucocidin, haemolysin.

LABORATORY DIAGNOSIS

SPECIMENS:

1- Sputum: microscopy, culture.

2- Blood for culture.

3- CSF: microscopy, culture, biochemistry.

MICROSCOPY:S.pneumoniae: @Gram positive, non-motile, diplococcus, capsulated.

@ Identified serologically (quelling reaction) using polyvalent antiserum.

CULTURE:

@ Cultivated on chocolate agar (10% CO2 ) @ Colonies flat with raised edges, giving a ring shape (draughtsman). @ α-haemolytic, & sensitive to optochin.

BIOCHEMICAL REACTIONS

@ Bile solubility positive .

ANTIMICROBIAL SENSITIVITY

@ Sensitive to: Penicillin, erythromycin, co-trimoxazole.

ANAEROBIC COCCI

SPECIES:

* Peptococcus * Peptostreptococcus.

HABITAT: skin, mouth, vagina, gastro-intestinal tract.

PATHOGENICITY:

1- Septicaemia, puerperal sepsis.

2- Bone and joint infections.

3- Abscesses, deep infected wounds, and ulcers.

MICROSCOPY:@ Gram positive:

•in chains: (Peptostreptococcus)

•in groups: (Peptococcus)

@ Catalase negative.

CULTURE:@ Cultivated in: Thioglycollate broth.

@ On B.A. colonies are small, shiny, non-haemolytic.

ANTIMICROBIAL SENSITIVITY

@ Sensitive to: Penicillin, metronidazole, @ Peptococci are resistant to novobiocin, @ Peptostreptococci are sensitive to novobiocin.

SpeciesSpeciesCatalasCatalasee

BacitraciBacitracinn

OptochiOptochin Bilen Bile

LitmuLitmus milks milk

CAMPCAMPEsculin Esculin hydrolyshydrolys

is is

S.pyogenes S.pyogenes (group A)(group A)--++--------

S.agalactiaS.agalactiaee

(group B)(group B)--------++--

E.faecalis E.faecalis (group D)(group D)------++--++S.viridansS.viridans------------

S.pneumoniS.pneumoniaeae----++------

FEATURES OF STREPTOCOCCI