Corynebacteria

Corynebacteria - Overview
 Gram positive, non motile bacilli with irregularly stained
segments

 Frequently show club shaped swellings – corynebacteria

(coryne = club)

 C. diphtheriae : most important member of this genus,

causes diphtheria

 Diphtheroids : commensals of nose, throat, nasopharynx,

skin, urinary tract & conjunctiva.
Introduction – C. diphtheriae
 Diphtheros – leather (tough, leathery
pseudomembrane)

 Also known as Klebs–Loeffler bacillus

 Causes Diphtheria
Important features of C. diphtheriae
 Slender Gram positive bacilli
 Pleomorphic, non motile, non sporing
 Chinese letter or Cuneiform arrangement
 Stains irregularly, tends to get easily
decolorised
 May show clubbing at one or both ends -
Polar bodies/ Metachromatic granules/
volutin or Babes Ernst granules
 Metachromatic Granules:
 made up of polymetaphosphate
 Bluish purple color with Loeffler’s
Methylene blue
 Special stains: Albert’s, Neisser’s &
Ponder’s
 Grows aerobically at 37°C
Virulence factor
 Exotoxin – Diphtheria toxin:
 Protein in nature
 very powerful toxin
 Responsible for all pathogenic
effects of the bacilli
 Produced by all the virulent
strains
 Two fragments A & B
 Disease is caused by exotoxin which
inhibits protein synthesis of eukaryotic
cells
 The toxin contains a toxic A subunit
(active toxin) and the receptor binding B
subunit. 
 The B subunit (fragment) facilitates
translocation of the A subunit from the
phagosome to the cytosol, followed by
separation, allowing full activity of the A subunit
on its target protein elongation factor-2.
 EF-2 transfers polypeptidyl-transfer RNA from
acceptor to donor sites on the ribosome of the
host cell. 
 The A subunit catalyzes transfer of
adenine ribose phosphate from NAD to
EF-2 (ADP ribosylation), inactivating EF-2,
and turning off protein synthesis, C.
diphtheria toxin is able to inhibit protein
synthesis of all eukaryotic cells. 
 Fragment A has the enzymatic activity
 Fragment B attaches to cellular
receptors and grants fragment A
entrance into the cell, where it inhibits
protein synthesis.
 The exotoxin is absorbed into the blood
demyelinating neuritis and myocarditis.
 The diphtheria toxin also causes local
destruction at the site of membrane
formation.
Diphtheria Toxin
 Toxigenicity can be induced by Lysogenic or phage
conversion – corynephages (tox+ phage) or beta
phages

 Can be toxoided by -
1. Prolonged storage
2. Incubation at 37°C for 4 - 6 weeks
3. Treatment with 0.2 – 0.4 % formalin or
4. Acid pH.

 Antibodies to fragment B - protective

Epidemiology
 Habitat – nose, throat, nasopharynx & skin of carriers and patients

 Spread by respiratory droplets, usually by convalescent or

asymptomatic carriers

 Nasal carriers harbour the bacilli for longer time than pharyngeal
carriers

 Local infection of throat - toxemia

 Incubation period of diphtheria – 3 to 4 days

 In tropics, cutaneous infection is more common than respiratory

infection
Diphtheria
 Site of infection
1. Faucial (palatine tonsil) – commonest type
2. Laryngeal
3. Nasal
4. Otitic
5. Conjunctival
6. Genital – vulval, vaginal, prepucial
7. Cutaneous – usually a secondary infection on pre-
existing lesion, caused by non toxigenic strains
Pathogenesis & Clinical Manifestations
 Human Disease
1. Usually begins in respiratory tract
2. Virulent diphtheria bacilli lodge in throat of
susceptible individual
3. Multiply in superficial layers of mucous
membrane
4. Elaborate toxin which causes necrosis of
neighboring tissue cells
5. Inflammatory response eventually results in
pseudomembrane (fibrinous exudate with
disintegrating epithelial cells, leucocytes,
erythrocytes & bacteria)

 Usually appears first on tonsils or posterior

pharynx and spreads upward or down
 In laryngeal diphtheria, mechanical
obstruction may cause suffocation
 Regional lymphnodes in neck often enlarged
(bull neck)
Diphtheria - Clinical Classification
 Based on the severity of clinical presentation:
1. Malignant or hypertoxic – severe toxemia with
marked adenitis

2. Septic – ulceration, cellulitis, & gangrene around the

pseudomembrane

3. Hemorrhagic – bleeding from the edge of

membrane, epistaxis, conjunctival hemorrahge,
purpura & generalized bleeding tendency.
Complications of diphtheria
 Mechanical complications are due to the
pseudomembrane, while the systemic effects
are due to the toxin.
1. Asphyxia – due to obstruction of respiratory
passage
2. Acute circulatory failure
3. Postdiphtheritic paralysis – occurs in 3rd or 4th week
of disease, palatine & ciliary, spontaneous
recovery
4. Sepsis – pneumonia & otitis media
Diphtheria Systemic complications
 Nerves
 toxicperipheral neuropathy
 paralysis of short nerves
 mouth, eye, facial extremities
 Cardiac
 Congestiveheart failure
 high amount of toxin 48-72 hours
 Low amount of toxin 2-6 weeks
 Laboratory Diagnosis
 Specimen – swab from the
lesions

1. Microscopy
 Gram stain: Gram +ve bacilli,
chinese letter pattern
 Immunofluorescence
 Albert’s stain for
metachromatic granules
Laboratory Diagnosis
2. Culture – isolation of bacilli requires media
enriched with blood, serum or egg
a. Blood agar
b. Loeffler’s serum slope – rapid growth, 6 to 8 hrs
c. Tellurite blood agar – tellurite is reduced to
tellurium, gives gray or black color to the colonies
Growth of diphtheria bacilli

Blood agar

Tellurite blood agar

Loeffler’s serum slope

Laboratory Diagnosis

3. Biochemical reactions
a. Hiss's serum water - ferments sugar with acid
formation but not Gas
ferments: glucose, galactose, maltose and dextrin

b. Resistant to light, desiccation and freezing

c. Sterilization: sensitive to heat (destroyed in 10mins

at 58°C or 1min in 100°C), chemical disinfectants
Laboratory Diagnosis
4. Virulence tests - Test for toxigenicity
A. Invivo tests – animal inoculation (guinea
pigs)
a. Subcutaneous test
b. Intracutaneous test

B. Invitro tests
a. Elek’s gel precipitation test
b. Tissue culture test
 Laboratory Diagnosis
Virulence tests - Invivo tests
 Bacterial growth from Loeffler’s serum slope is emulsified in 2-4 ml
broth.
 Two guinea pigs (GP A and GP B)

I. Subcutaneous test – 0.1 ml of emulsion is injected SC into each

guinea pig
GP A - has diphtheria antitoxin (500 units injected 18 to 24 hours before)
GP B - Doesn't have antitoxin

II. Intracutaneous test - 0.1 ml of emulsion is injected IC into each

guinea pig
GP A - has diphtheria antitoxin (500 units injected 18 to 24 hours before)
GP B – 50 units of antitoxin IP four hrs after the skin test
 Laboratory Diagnosis
Virulence tests - Invitro tests
I. Elek's gel precipitation test
 filter paper saturated with antitoxin (1000units/ ml) is placed on
agar plate with 20% horse serum
 bacterial culture streaked at right angles to filter paper
 Laboratory Diagnosis
Virulence tests - Invitro tests
II. Tissue culture test
- incorporation of bacteria into agar overlay of
eukaryotic cell culture monolayers.

Result: toxin diffuses into cells and kills them

Prophylaxis
1) Active Immunization (Vaccination)
Prophylaxis
 Adsorbed Toxoid
 SHICK test - to test susceptibility to vaccine, not done now-a-days
Prophylaxis
2. Passive immunization
Combined immunization
CONTROL
1. isolate patients
2. treat with antibiotics actively
3. complete vaccination schedule should be
used with booster every 5 years