Streptococci microbiology information...

Microbiology

1. StreptococciStreptococci
By
Dr Moeen Zafar
Visiting Lecturer pathology
IPMR KMU
Peshawar

2. StreptococciStreptococci
Characteristics of Streptococci
oGram positive cocci
o1µm in diameter
oChains or pairs
oUsually capsulated
oNon motile
oNon spore forming
oFacultative anaerobes
oCatalase negative (Staphylococci are catalase positive)

4. Classification of StreptococciClassification of Streptococci
Streptococci can be classified according to:
A. Oxygen requirements
•Anaerobic (Peptostreptococcus)
•Aerobic or facultative anaerobic (Streptococcus)
B. Serology (Lancefield Classification)
C. Hemolysis on Blood Agar (BA)

5. Serology: Lancefield ClassificationSerology: Lancefield Classification
•Rebecca Lancefield developed useful serogrouping system
•Streptococci classified into many groups from A-U
•One or more species per group
•Classification based on C- carbohydrate antigen of cell wall

7. Classification of Streptococci Based on Hemolysis
on Blood Agar
Hemolysis on BA
-hemolysis
Partial hemolysis
Green discoloration around the colonies
e.g. non-groupable streptococci (S. pneumoniae & S. viridans)
-hemolysis
Complete hemolysis
Clear zone of hemolysis around the colonies
e.g. Group A & B (S. pyogenes & S. agalactiae)
-hemolysis
No lysis
e.g. Group D (Enterococcus spp)

8. Hemolysis on Blood agar
-hemolysis
-hemolysis
-hemolysis

9. Group A streptococciGroup A streptococci
•Include only S. pyogenes
•Group A streptococcal infections affect all ages peak incidence at 5-15
years of age
•90% of cases of pharyngitis
•Also common cause of skin infections
•Growth of S. pyogenes is inhibited by
antibiotic bacitracin……important
diagnostic criterion.

10. Antigenic Structure
Streptococcus pyogenes (Group A)
1. C carbohydrate
oGroup specific antigen
odetermines the group of Beta hemolytic streptococci.
oIt is located in the cell wall
2. M protein
oType specific antigen
oAntibody to M protein provides type-specific immunity
o There are approximately 80 serotypes based on the M protein
odetermines the type of group A Beta hemolytic streptococci.
omost important virulence factor
o Antiphagocytic.

11. Antigenic StructureAntigenic StructureCont……..Cont……..
3. Polysaccharide Capsule
oComposed of Hyaluronic acid
oSame as host hyaluronic acid (cartilage, skin etc)
oAntiphagocytic
oAntibodies are not formed against the capsule because hyaluronic acid is
a normal component of the body and humans are tolerant to it.
4. T protein: type-specific; function unknown.
5. Protein F: a major adhesin of S. pyogenes
6. Lipoteichoic acid: adhesion

12. Pathogenesis of S. pyogenesPathogenesis of S. pyogenes
Group A streptococci (Str. pyogenes) cause disease by three mechanisms:
(1) Pyogenic inflammation:
o which is induced locally at the site of the organisms in tissue.
(2) Exotoxin production:
ocause widespread systemic symptoms in areas of the body where there are
no organisms
(3) Immunologic:
owhich occurs when antibody against a component of the organism cross-
reacts with normal tissue or forms immune complexes that damage normal
tissue.
oThe immunologic reactions cause inflammation, e.g. , the inflamed joints of
rheumatic fever , but there are no organisms in the lesions

13. Pathogenesis and Virulence FactorsPathogenesis and Virulence Factors
Enzymes
1. Streptokinase (fibrinolysin):
Activate plasminogen to form plasmin dissolve fibrin in clot, thrombi and emboli.
Used to lyse thrombi in arteries of MI patients.
2. Streptodernase (DNase A to D) :
oDegrades DNA in exudates or necrotic tissue
oA mixture of Streptodernase and streptokinase is used for enzymatic
liquefaction of exudates and facilitates removal of pus and necrotic tissue.
3. Hyaluronidase (spreading factor):
odegrades hyaluronic acid, which is the ground substance of subcutaneous tissue.
oHyaluronidase is known as spreading factor because it facilitates the rapid spread of Str.
pyogenes in skin infections (cellulitis).

14. Toxins and hemolysins:
•Erythrogenic toxin:
oCauses the rash of scarlet fever
oErythrogenic toxins are known to damage the plasma membranes of blood
capillaries under the skin and produce a red skin rash (characteristic of scarlet
fever)
oIts mechanism of action is similar to that of the toxic shock syndrome toxin (TSST)
of Sta. aureus, i.e. , it acts as a superantigen
oIt is produced only by certain strains of Str.pyogenes lysogenized by a
bacteriophage carrying the gene for the toxin

15. Toxins and hemolysins Cont……….
•Pyogenic exotoxin A:
oResponsible for most cases of streptococcal toxic shock syndrome
oIt has the same mode of action as does staphylococcal TSST , i.e. , it is a
super antigen that causes the release of large amounts of cytokines from
helper T cells and macrophages
•Exotoxin B:
ois a protease that rapidly destroys tissue and is produced in large amounts
by the strains of Str. pyogenes, the so-called "flesh eating" streptococci
that cause necrotizing fasciitis.

16. Toxins and hemolysins
Streptolysins
a. Streptolysin O
ois a hemolysin
olyse red blood cells, white blood cells, and platelets
oinactivated by oxidation (oxygen-labile).
oIt causes hemolysis only when colonies grow under the surface of a blood agar
plate.
oIt is antigenic, and antibody to it (ASO) develops after group A streptococcal
infections.
oThe titer of ASO antibody can be important in the diagnosis of rheumatic fever .

17. Streptolysins
b. Streptolysin S
ois a hemolysin
oNot inactivated by oxygen (oxygen-stable).
oIt is not antigenic but is responsible for hemolysis when
colonies grow on the surface of a blood agar plate.

18. Disease caused by S. pyogenes
Str. pyogenes causes three types of diseases:
(1)Pyogenic diseases such as pharyngitis and cellulitis,
(2)Toxigenic diseases such as scarlet fever and toxic shock syndrome, and
(3)Immunologic diseases such as rheumatic fever and acute
glomerulonephritis (AGN)

19. Disease caused by S. pyogenes
Suppurative
1.Sore Throat (Pharyngitis)
omost common bacterial cause of sore throat.
oPharyngitis (“strep throat”)-inflammation of the
Pharynx
oPharyngitis is characterized by inflammation, exudate, fever , leukocytosis, and
tender cervical lymph nodes
oIf untreated, spontaneous recovery occurs in 10 days, but rheumatic fever may
occur
oUntreated pharyngitis may extend to the middle ear (otitis media), the sinuses
(sinusitis), the mastoids (mastoiditis), or the meninges (meningitis)

20. Disease caused by S. pyogenes
2. Skin and Soft tissues infections:
oGroup A streptococci cause skin and soft tissue infections, such as
Cellulitis,
Erysipelas (large raised red patches on the skin),
Necrotizing fasciitis (streptococcal gangrene),
and impetigo.
Necrotizing fasciitisErysipelas Scarlet fever-rash
Cellulitis

21. Skin and Soft tissues infectionsCont…
oImpetigo, a form of pyoderma, is a superficial skin infection characterized
by "honey-colored“ crusted lesions.
oLymphangitis can occur , especially on the forearm associated with an
infection on the hand
oScarlet fever-
orash that begins on the chest and spreads across the body
o Strawberry tongue, fever, sore throat
Lymphangitis

22. Post streptococcal (Nonsuppurative) Diseases
oThese are disorders in which a local infection with group A streptococci is
followed weeks later by inflammation in an organ that was not infected by
the streptococci.
oThe inflammation is caused by an immunologic (antibody) response to
streptococcal M proteins that cross-react with human tissues.
oSome strains of Str. pyogenes bearing certain M proteins are nephritogenic
and cause AGN,
oand other strains bearing different M proteins are rheumatogenic and cause
acute rheumatic fever .
oNote that these diseases appear several weeks after the actual infection
because that's the length of time it takes to produce sufficient antibodies.

23. Poststreptococcal (Nonsuppurative) Diseases
Cont……………….
Rheumatic fever:
oLife threatening inflammatory disease that leads to damage of heart valves
muscle
oApproximately 2 weeks after a group A streptococcal infection— usually
pharyngitis —rheumatic fever
ocharacterized by fever ,migratory polyarthritis, and carditis may develop.
oThe carditis damages myocardial and endocardial tissue, especially the mitral and
aortic valves
oUncontrollable, spasmodic movements of the limbs or face (chorea) may also
occur .
oASO titers and the erythrocyte sedimentation rate are elevated

24. Rheumatic fever Cont……
oNote that group A streptococcal skin infections do not cause rheumatic
fever .
oMost cases of pharyngitis caused by group A streptococci occur in children
aged 5 to 15 years and hence rheumatic fever occurs in that age group
oRheumatic fever is due to an immunologic reaction between cross-reacting
antibodies to certain streptococcal M proteins and antigens of joint, heart,
and brain tissue.
oIt is an autoimmune disease, greatly exacerbated by recurrence of
streptococcal infections.
oIf streptococcal infections are treated within 8 days of onset, rheumatic
fever is usually prevented.
oIn the United States, fewer than 0.5% of group A streptococcal infections
lead to rheumatic fever , but in developing tropical countries, the rate is
higher than 5%

25. •ACUTE GLOMERULONEPHRITIS
oAGN typically occurs 2 to 3 weeks after skin infection by certain group A
streptococcal types in children (e.g. , M protein type 49 causes AGN most
frequently).
oAGN is more frequent after skin infections than after pharyngitis.
oThe most striking clinical features are:
Hypertension,
Edema of the face and ankles, and
“Smoky" urine (due to red cells in the urine).
oMost patients recover completely. Reinfection with streptococci rarely leads to
recurrence of glomerulonephritis.
oIt can be prevented by early eradication of nephritogenic streptococci from skin
colonization sites but not by administration of penicillin after the onset of
symptoms.

26. Group B streptococci (Str. Agalactiae):
oColonize vagina
ocause neonatal sepsis and meningitis.
oAlso an important cause of neonatal pneumonia
oAlthough most group B streptococcal infections are in neonates, this
organism also causes such infections as pneumonia, endocarditis, arthritis, and
osteomyelitis in adults.
oPostpartum endometritis also occurs.
oDiabetes is the main predisposing factor for adult group B streptococcal
infections.
oBacitracin resistant

27. Group D streptococci
• Enterococci (e.g. , Enterococcus faecalis and Enterococcus faecium) and
nonenterococci (e.g. , Str. bovis)
Enterococci
•members of the normal flora of the colon
•cause urinary, biliary, and cardiovascular infections
•Indwelling urinary catheters and urinary tract instrumentation are important
predisposing factors
•also cause endocarditis, particularly in patients who have undergone gastrointestinal or
urinary tract surgery or instrumentation
•They are very hardy organisms
•can grow in hypertonic (6.5%) saline or in bile
•Not killed by penicillin G.
•Combination of penicillin and aminoglycoside is needed. Or vancomycin.

28. Cont….…..Cont….…..
Nonenterococcal group D streptococci, such as Str. bovis,
•can cause similar infections
•causes endocarditis, especially in patients with carcinoma of the
colon.
•but are much less hardy organisms, e.g. ,
•they are inhibited by 6.5% NaCl and killed by penicillin G

29. Non beta hemolytic Streptococci
• Str. pneumoniae and the viridans group of streptococci
• Alpha hemolytic organisms
•Viridans streptococci
•Species include, such as Str. sanguis, Str. mutans, Str. mitis, Str. gordoni,
Str. salivarius, Str. anginosus, Str. milleri, and Str. intermedius.
• Viridans streptococci are part of the normal flora of the human pharynx
and intermittently reach the bloodstream to cause infective endocarditis.
Viridans streptococci Str. pneumoniae
Bile-solubility Soluble Not soluble
Optochin sensitivityResistant Sensitivity

30. Non beta hemolytic Streptococci
Cont….......
•Str. mutans synthesizes polysaccharides (dextrans) that are found in dental
plaque and lead to dental caries.
•They are found primarily in the mouth and colon
Viridans streptococci (e.g. , Str. mutans, Str. sanguis, Str. salivarius, and Str.
mitis) are the most common cause of infective endocarditis.
•Dental surgery------blood stream (bacteremia)------Endocarditis
Viridans streptococci, especially Str. anginosus, Str. milleri, and Str.
intermedius, also cause brain abscesses, often in combination with mouth
anaerobes (a mixed aerobic–anaerobic infection)
• Dental surgery-----viridans streptococci and the anaerobes----- enter the
bloodstream (bacteremia)-----spread to the brain

31. Laboratory DiagnosisLaboratory Diagnosis
Specimen collection:
Throat swab,
Pus or
Blood
depending upon the site
oCultures of swabs from the pharynx or lesion on blood agar plates show small,
translucent, Beta hemolytic colonies in 18 to 48 hours.

32. Differentiation between Differentiation between -hemolytic streptococci-hemolytic streptococci
1. Bacitracin susceptibility Test:
odifferentiate between -hemolytic streptococci
oSpecific for S. pyogenes (Group A)
oIf the growth is inhibited by bacitracin disk, they are likely to be group A
streptococci

33. Laboratory DiagnosisLaboratory Diagnosis
2. CAMP test (Christie,
 Atkins, and Munch-Peterson):
oSpecific for S. agalactiae (Group B)
oPositive for S. agalactiae (Group B)

34. Differentiation between Viridans group streptococci and Str. pneumoniae
oBoth are Alpha hemolytic
HemolysisOptochin
sensitivity
Bile
solubility
Inulin
Fermentation
S. pneumoniae  Sensitive (≥
14 mm)
Soluble Not ferment
Viridans strep  Resistant
(≤13 mm)
Insoluble Ferment
Differentiation between -hemolytic streptococci

35. Optochin Susceptibility TestOptochin Susceptibility Test
Optochin susceptible
S. pneumoniae
Optochin resistant
S. viridans

36. •Bile Solubility test
oS. pneumoniae soluble in bile whereas S. viridans
insoluble

37. Differentiation between -hemolytic streptococci
Hemolysis Bacitracin
sensitivity
CAMP test
S. pyogenes  Susceptible Negative
S. agalactiae  Resistant Positive
HemolysisOptochin
sensitivity
Bile
solubility
Inulin
Fermentation
S. pneumoniae  Sensitive (≥
14 mm)
Soluble Not ferment
Viridans strep  Resistant
(≤13 mm)
Insoluble Ferment
Differentiation between -hemolytic streptococci

38. Streptococcus pneumoniae
•General Characteristics
oPneumococci are gram-positive
olancet-shaped cocci oval
oarranged in pairs (diplococci) or short chains
o On blood agar they produce alpha hemolysis.
oCapsulated
oBile soluble
oOptochin Sensitive
oCatalase Negative
o Possesses a large capsule to resist phagocytosis
o>85 different serotypes based on capsule protein as Ag.
oURT resident

39. Pathogenicity and virulence
1. Capsule
•More than 90 antigenically distinct types based on the nature of capsular
polysaccharides.
•quelling reaction (type specific antiserum,capsule swells)
2. IgA Protease : cleave IgA …consolidation of lung
3. Cell wall Carbohydrate C substance:
•Present in all pneumococci
•Species specific carbohydrate antigen
4. Toxins – pneumolysin
•Membrane damaging toxin
•Cytotoxic and complement activating property

40. PathogenicityPathogenicity
Source of infection:
i)Endogenous: from the colonized area.
ii)Exogenous: patients or carriers.
Mode of infection: By inhalation.

41. Mechanism of PathogenesisMechanism of Pathogenesis::
Entry of pneumococci into nasopharynx
Colonization of nasopharynx
May cause infection of the middle ear, Para nasal
sinuses & respiratory tract by direct spread
Infection of meninges can also occur, through blood
Enters blood causing bacteremia, which may also
lead to disseminated infections as in the heart,
peritoneum or joint

42. DISEASES CAUSED BY STREPTOCOCCUS PNEUMONIAE
Non-invasive disease
•Pneumonia (lungs)
•Sinusitis (sinuses)
•Otitis media (middle ear)
Invasive disease
•Bacteremia (blood)
•Meningitis (CNS)
•Endocarditis (heart)
•Peritonitis (body cavity)
•Septic arthritis (bones and joints
•Others (appendicitis, salpingitis, soft-tissue infections)

43. MAIN SYMPTOMS OF INFECTION
Clinical Findings
•Pneumonia often begins with a sudden chill, fever, cough, and pleuritic pain.
•Sputum is a red or brown “rusty” color.
•Bacteremia occurs in 15% to 25% of cases.
•Spontaneous recovery may begin in 5 to 10 days
•Pneumococci are a prominent cause of otitis media, sinusitis, mastoiditis,
conjunctivitis, purulent bronchitis, pericarditis, bacterial meningitis, and sepsis.
•Pneumococci are the leading cause of sepsis in patients without a functional
spleen.

44. LABORATORY DIAGNOSIS:LABORATORY DIAGNOSIS:
Specimens to be collected:
•Sputum,
•CSF,
•Blood,
•Synovial fluid,
•In children laryngeal swab can be taken if sputum can not be
collected

45. LABORATORY DIAGNOSIS Cont……
Methods of examination:
1.Direct microscopy:
Gram stained
smears reveals
Gram positive
lancet shaped
diplococci with
numerous pus cells.

46. LABORATORY DIAGNOSIS Cont……
•On blood agar , pneumococci form small Alpha hemolytic colonies.
•The colonies are bile-soluble, i.e. , are lysed by bile, and growth is inhibited
by optochin
2. Quelling( capsular swelling ) reaction:
•It is described by Neufeld.
•On a slide the sputum is mixed with type specific antiserum against capsular
antigen & a loop ful of methylene blue solution.
•The capsule becomes swollen.

47. Summary Summary
Streptococci
Gram positive cocci in Chains or pairs
Facultative anaerobes
Catalase negative
Classification of
Streptococci
Hemolysis on BA
-hemolysis
Partial hemolysis
Green discoloration around the colonies
e.g. non-groupable streptococci (S. pneumoniae &
S. viridans)
-hemolysis
Complete hemolysis
Clear zone of hemolysis around the colonies
e.g. Group A & B (S. pyogenes & S. agalactiae)
-hemolysis
No lysis
e.g. Group D (Enterococcus spp)
Oxygen requirements
Anaerobic (Peptostreptococcus)
Aerobic or facultative anaerobic
(Streptococcus)
Serology(Lancefield Classification)
Hemolysis on Blood Agar (BA)

48. Group A streptococci
S. pyogenes
•peak incidence at 5-15 years of age
•90% of cases of pharyngitis
•common cause of skin infections
•Bacitracin sensitive
Antigenic Structure
Streptococcus pyogenes (Group A)
C carbohydrate
oGroup specific antigen
M protein
oType specific antigen
omost important virulence factor
o Antiphagocytic.
Polysaccharide Capsule
oComposed of Hyaluronic acid
oAntiphagocytic
Lipoteichoic acid:
oadhesion
•Protein F
•T protein
Pathogenesis of S. pyogenes
(1) Pyogenic inflammation:
oinduced locally at the site of the organisms in tissue.
(2) Exotoxin production:
ocause widespread systemic symptoms in areas of the body
where there are no organisms
(3) Immunologic:
owhich occurs when antibody against a component of the
organism cross-reacts with normal tissue or forms immune
complexes that damage normal tissue.
oThe immunologic reactions cause inflammation, e.g. , the
inflamed joints of rheumatic fever , but there are no
organisms in the lesions

49. Pathogenesis and Virulence FactorsPathogenesis and Virulence Factors
Enzymes
Streptokinase (fibrinolysin):
Activate plasminogen to form plasmin dissolve
fibrin in clot, thrombi and emboli.
Used to lyse thrombi in arteries of MI patients.
Streptodernase (DNase A to D) :
Degrades DNA in exudates or necrotic tissue
A mixture of Streptodernase and streptokinase is
used for enzymatic liquefaction of exudates and
facilitates removal of pus and necrotic tissue.
Hyaluronidase (spreading factor):
degrades hyaluronic acid
it facilitates the rapid spread of Str. pyogenes
in skin infections (cellulitis).
Toxins and hemolysins::
•Erythrogenic toxin:
oCauses the rash of scarlet fever
•Pyogenic exotoxin A:
oResponsible for streptococcal toxic shock syndrome
•Exotoxin B:
ois a protease that rapidly destroys tissue
o"flesh eating" streptococci that cause necrotizing
fasciitis.
a. Streptolysin O
ois a hemolysin, oxygen-labile
olyse RBC’s, WBC’s, and platelets
odiagnosis of rheumatic fever .
b. Streptolysin S
ois a hemolysin, oxygen-stable, not antigenic

50. Disease caused by S. pyogenes
Str. pyogenes causes three types of
diseases:
Pyogenic diseases such as
pharyngitis and cellulitis.
Pharyngitis
oIf untreated, spontaneous recovery occurs
in 10 days, but rheumatic fever may occur
oUntreated pharyngitis may extend to the
middle ear (otitis media), the sinuses
(sinusitis), the mastoids (mastoiditis), or the
meninges (meningitis)
Skin and Soft tissues infections:
•Cellulitis, erysipelas, necrotizing fasciitis,
impetigo, Lymphangitis, Scarlet fever-rash
Toxigenic diseases such as
scarlet fever and toxic shock
syndrome.
Immunologic diseases such as
rheumatic fever and acute
glomerulonephritis (AGN)

51. Group B streptococci (Str. Agalactiae):
oColonize vagina
ocause neonatal sepsis and meningitis
oneonatal pneumonia
Non beta hemolytic Streptococci
o Str. pneumoniae and the viridans group of
streptococci
oAlpha hemolytic organisms
Viridans streptococci
•Species include, such as Str. sanguis, Str. mutans,
Str. mitis, Str. gordoni, Str. salivarius, Str. anginosus,
Str. milleri, and Str. Intermedius
•Dental Caries
•Endocarditis
•Brain abscesses, often in combination with mouth
anaerobes (a mixed aerobic–anaerobic infection)
Group D streptococci
•Enterococci (e.g. , Enterococcus faecalis
and Enterococcus faecium) and
nonenterococci (e.g. , Str. bovis)
Enterococci
omembers of the normal flora of the
colon
ocause urinary, biliary, and cardiovascular
infections
Nonenterococcal group D streptococci,
such as Str. bovis,
•can cause similar infections
•causes endocarditis.

52. Streptococcus pneumoniae
•General Characteristics
oPneumococci are gram-positive
oarranged in pairs (diplococci) or
short chains
oCapsulated
oOptochin Sensitive
oBile soluble
oCatalase Negative
DISEASES
Non-invasive disease
•Pneumonia (lungs)
•Sinusitis (sinuses)
•Otitis media (middle ear)
Invasive disease
• Bacteremia (blood)
• Meningitis (CNS)
• Endocarditis (heart)
• Peritonitis (body cavity)
• Septic arthritis (bones and joints
• Others (appendicitis, salpingitis, soft-
tissue infections)

53. HemolysisOptochin
sensitivity
Bile
solubility
Inulin
Fermentation
S. pneumoniae  Sensitive (≥
14 mm)
Soluble Not ferment
Viridans strep  Resistant
(≤13 mm)
Insoluble Ferment
Differentiation between -hemolytic streptococci
HemolysisBacitracin sensitivity CAMP test
S. pyogenes  Susceptible Negative
S. agalactiae  Resistant Positive
Differentiation between Differentiation between -hemolytic streptococci-hemolytic streptococci

54. •References :
•Review of microbiology by Levinson
•Sketchy microbiology
•Lecture of Sir Ikhlaq lecturer MLT ,NWIHS.

55. THANK YOUTHANK YOU