mORPHOLOGY, PATHOGENESIS & LAB DIAGNOSIS OF STREPTOCOCCI

1. STREPTOCOCCISTREPTOCOCCI
Dr V S Vatkar
Asso Prof
Microbiology Department

2. DefinitionDefinition
• Gram position cocci
• in chains,
• non sporing,
• non motile,
• some capsulated,
• facultatively anaerobic and fastidious in
nutritional requirements.

3. IntroductionIntroduction
• Billroth (1874): in erysipelas & calledin erysipelas & called
them streptococcithem streptococci (streptos = twisted or(streptos = twisted or
coiled)coiled)..
• Ogston (1881) Isolated them from acuteIsolated them from acute
abscesses , differs from staphylococci &abscesses , differs from staphylococci &
established their pathogenicity by animalestablished their pathogenicity by animal
inoculation.inoculation.
• Rosenbach (1884) Isolated the cocci fromIsolated the cocci from
human suppurative lesions, named themhuman suppurative lesions, named them
Streptococcus pyogenesStreptococcus pyogenes..

4. Classification of Streptococci
Aerobes & Facultative anaerobes Obligate anaerobes
Peptostreptococcus
Oxygen requirement
hemolysis
α hemolysis
Streptococcus viridans
Pneumococci
β hemolysis
β hemolytic streptococci
γ hemolysis
Enterococci
Lancefield grouping
On the basis of C Ag
A-V except I & J
Gr A Streptococci
GRIFFITH CLASSIFICATION
Based on M protein & emm gene

5. Based on Haemolysis on BloodBased on Haemolysis on Blood
AgarAgar
(i) β-haemolytic Streptococci (BHS) – complete
haemolysis of the red cells around the colonies,
producing clear zones around them.
e.g. group A, group B etc…
(ιι) α-haemolytic Streptococci – partial haemolysis with
greenish discoloration of the areas surrounding the
colonies.
e.g.Streptococcus viridans, Streptococcus pneumoniae
(iii) Non-haemolytic Streptococci
e.g. Enterococcus faecalis

6. Lancefield GroupingLancefield Grouping
• Usually done on β-haemolytic streptococci
(BHS). Based on the presence of a
carbohydrate component of cell wall the C
carbohydrate. About 20 Lancefield groups
designated as A,B,C,D, (A-H) (K-U).
• Detected by reacting extract of carbohydrate C
antigen with specific antisera raised against it.

7. Alfa hemolysis
Beta hemolsis

8. M SerotypingM Serotyping
• Done on only group A streptococci and based
on the M protein found in Group A
Streptococci. 60 such serotypes; useful for
epidemiological studies.

9. Group A StreptococciGroup A Streptococci
(Lancefield Grouping)(Lancefield Grouping)
((Streptococci PyogenesStreptococci Pyogenes))
Morphology
• Spherical or oval
• In chains
• Nonmotile and nonsporing
• It may be capsulated and the capsule is
composed of hyaluronic acid.

10. CultureCulture
• Aerobes and facultative anaerobes
• 22°C - 42°C
• Colony characters: virulent strains: mat finish
colonies, avirulent strains:glossy colonies,
capsulated colonies : mucoid

11. Growth & Clonial MorphologyGrowth & Clonial Morphology
• Blood agar best medium with optimum
temperature of 35-37°C & under aerobic
conditions.
• Colonies after 24 hours incubation: about 0.5 –
1mm in diameter & may/may not be
surrounded by haemolysis.
• They are catalase negative.

12. ΒΒ hemolysis on blood agarhemolysis on blood agar

13. Biochemical reactionsBiochemical reactions
• Ferment various sugars
• Produce acid but not gas
• Not soluble in 10% bile
• PYR

14. ResistanceResistance
• Delicate organism
• Can be stored in RCM at 4°C
• More resistance to crystal violet
• Sensitive to bacitracin

15. Antigenic structure –Antigenic structure –
Virulence factorsVirulence factors
• Cell wall antigens composed of 3 layers
• Inner thick peptidoglycan layer: rigidity to cell
wall, induces inflammatory response,
thrombolytic activity
• Middle layer: C or carbohydrate Ag
• Outer layer: lipoteichoic acid layer: composed
of M,T and R proteins
• M protein : principle virulence factor, inhibits
phagocytosis, inflammatory response

17. ToxinsToxins
Streptolysin O
• Oxygen labile
• Soluble in oxygen so
named as Sreptolysin O
• Seen in deep colonies
• Strongly Agenic
• ASO : raised in many
streptococcal infections
• Increases in RF &
decreases in GN and
pyoderma
Streptolysin S
• Oxygen stable
• Serum soluble so named
streptolysin S
• Hemolysis on surface of
blood agar
• Not Agenic
• Not useful for serological
diagnosis

18. Erythrogenic or pyrogenic exotoxinErythrogenic or pyrogenic exotoxin::
streptococcal pyrogenic exotoxinstreptococcal pyrogenic exotoxin (SPE)(SPE)
• Scarlet fever, necrotizing fasciitis & TSS
• 3 Agenic subtypes: SPE A , B & C
• Subtype A & C : bacteriophage mediated , act
as super Ag ----- stimulation of T-cells ------
release of cytokines ----- fever, shock, tissue
damage
• SPE B: chromosomally mediated
• DICK test: I/D inj in susceptible children
produce local erythema. Initially uesd to detect
scarlet fever

19. EnzymesEnzymes
 Streptokinase: Convert plasminogen to
plasmin which then lyses fibrin. Used to treat
thrombotic states. e.g. Coronary thrombosis ,
MI
 Deoxyribonucleases
(Streptodornase):
4 main DNAases: A B C D
Antibodies produced against DNAase
(anti-DNAase B) is useful for diagnosing recent
Group A Streptococcal infections especially
skin infections.

20. • Di-phospho-pyridine-nucleotidase (DPN-ase)
 Hyaluronidase :Degrades hyaluronic acid
 Spy CEP: serine protease
 C5a peptidase

21. PathogenesisPathogenesis
•Causes suppurative infections and
non-suppurative complications (or
sequalae).

22. A. Suppurative (Pyogenic) Infections
a) Virulence Factors
(i) Principal virulence factors is the M protein
Originated from the cytoplasmic membrane.
Associated with pili.
It is antiphagocytic.
(ii) Lipotechoic Acid (LTA)
For attachment to epithelial surfaces.
(iii) Hyaluronic Acid
An antiphagocytic capsules.

23. pathogenesity

24. PathogenicityPathogenicity
 Respiratory tract
 Acute tonsillitis
 Pharyngitis
 Scarlet fever
 Otitis media, Mastoiditis,
 Quinsy, Ludwig's angina
 Skin infections
 Wound infections, Burns, Cellulitis,
 Erysipelas, Impetigo or Pyoderma : An infection
of the epidermis presenting as pustules. Seen
most often in infants and toddlers.

25. Strawberry tongue in scarlet fever
cellulitis erysipelas

26. SCARLET FEVERSCARLET FEVER
DWDDWDDesquamationDesquamation

27. Flesh Eating BacteriaFlesh Eating Bacteria
• Cellulitis- leading toCellulitis- leading to
Necrotising FasciitisNecrotising Fasciitis
• Leads to necrosis ofLeads to necrosis of
subcutaneoussubcutaneous
issues andissues and
muscular tissuesmuscular tissues
and adjutant fasciaand adjutant fascia
• Other complicationsOther complications
Toxic shockToxic shock
syndromesyndrome

28. INVASIVE GROUP A STREPTOCOCCI AND “FLESH-INVASIVE GROUP A STREPTOCOCCI AND “FLESH-
EATING” SYNDROME OR NECROTIZING FASCIITISEATING” SYNDROME OR NECROTIZING FASCIITIS
Caused by virulent strainsCaused by virulent strains
ofof StreptococcusStreptococcus
pyogenespyogenes

29. PathogenicityPathogenicity
• Genital infections Both aerobic & anaerobic are normal
inhabitants of female genitalia
• Deep infections Bone & joint infections
Lymphadenitis, Septicaemia
• Non suppurative complications: Ac. rheumatic
fever & Ac. Glomerulonephritis
These are antigen-antibody mediated disease and occur about 1-5
weeks after the primary suppurative infection. Tend to follow
either throat or skin infections or both. Streptococci are not
found in the affected organ.

30. NON SUPPURATIVE LESIONSNON SUPPURATIVE LESIONS
• Acute RheumaticAcute Rheumatic
FeverFever
• AcuteAcute
glomerulonephritisglomerulonephritis
• Carditis.Carditis.
• Involvement ofInvolvement of
Connective tissuesConnective tissues
of the heartof the heart

31. NON SUPPURATIVE LESIONSNON SUPPURATIVE LESIONS

32. a)a) Acute Rheumatic Fever:Acute Rheumatic Fever:
• Considered to be an autoimmune disease involving the
myocardium and its valves, connective tissues and the
big joints.
• Group A Strep cell wall has some antigenic similarity
with some of these human tissues. Follows after throat
infections only. Tends to recur. Many serotypes are
associated with acute rheumatic fever.

34. b)b)Acute Glomerulonephritis:Acute Glomerulonephritis:
• Due to antigen-antibody complexes deposited on the
basal membrane of glomeruli also can be due to
similarity between group A cell components and
glomerular tissue. May follow after either throat or skin.
Tends not to recur. Serotypes involved are few called
nephrotogenic strains.

35. Differences Between Glomerulonephritis &Differences Between Glomerulonephritis &
Rheumatic FeverRheumatic Fever
1. Latent period between
infection and first attack.
1 – 5 weeks
(Average 18 days)
1 – 5 weeks
(Average 10 days)
2. Preceding infection Throat only Throat or Skin
3. Pathogenesis Both Based On
Immunological Reaction
(Either Due to auto antibody
Or due to cross reactive
antigen).
Similarity between
organism antigens
& tissue antigens
Similarity between
a) Organism & tissue
antigens.
b) Deposition of
immunocomplexes
in glomeruli
Rheumatic Fever Glomerulonephritis

36. 4. Second Attacks Common Rare if any
5. Prophylactic use of penicillin. Essential Usually NOT used.
6. Serotypes (M Types) Any of the 60
serotypes
Limited No. of
serotypes e.g.
type 12, 45 etc.
7. Serum whole complement &
C3
Increased Decreased
Rheumatic Fever Glomerulonephritis
Differences Between Glomerulonephritis &Differences Between Glomerulonephritis &
Rheumatic Fever (Continued)Rheumatic Fever (Continued)

37. Epidemiology ofEpidemiology of StreptococcalStreptococcal InfectionsInfections
1. Infection acquired through infected respiratory
droplets.
2. Sources of Infection
a) Those with active disease or convalescent carriers in
throat.
b) Asymptomatic carriers – the most common source. Up to
20% of school going children may carry Group A
streptococci in their throats.
3. Age Group: prevalent in children especially
between 3 – 8 years.

38. Lab. DiagnosisLab. Diagnosis
A] In acute infections : by culture
• Specimen : swab,pus,blood, CSF
• Collection & transport
• Culture

39. Lab. DiagnosisLab. Diagnosis
• Biochem.tests
. Catalase negative
. Sugar fermentation with production of
acid but not gas
. PYR hydrolysis
. Fluorescent antibody technique
. Bacitracin test

40. Lab. DiagnosisLab. Diagnosis
. Anti-deoxy-ribonuclease B (ADN-ase)
. Streptozyme test
. Lancefield grouping
Todd hewitt broth
Fullers method
Rantz & Randall’s method
Maxted, method
. Antigen detection test : by ELISA

41. ProphylaxisProphylaxis
Indicated only in prevention of Rh.fever
to prevent reinfection and damage to the
heart
Long term Penicillin is given to the child
who develops early stages of Rh.fever

42. TreatmentTreatment
• Drug of choice is Penicillin
• Erythromycin
• Cephalexin
• Tetracycline's & sulphonamides are not
recommended
• Chloramphenicol
• Marolides

43. THANK YOUTHANK YOU

44. Gram stain & col.on Blood agarGram stain & col.on Blood agar

45. Cell surface with secreted productsCell surface with secreted products

46. PathogenesisPathogenesis

47. Phagocytosis of str.pyogenes by aPhagocytosis of str.pyogenes by a
macrophagemacrophage

48. Dividing streptococci 12000 xDividing streptococci 12000 x

49. Negative staining of group A streptococci viewed by TEMNegative staining of group A streptococci viewed by TEM
28,000X. The "halo" around the chain of cells (approximately28,000X. The "halo" around the chain of cells (approximately
equal in thickness to the cell diameter) is the remnants of theequal in thickness to the cell diameter) is the remnants of the
capsulecapsule