Transcript Mycobacterium tuberculosis

Corynebacterium diphtheriae
Morphology
•Another characteristic is the presence of metachromatic granules
(RNA and polymetaphosphate) in bacterial cells. The granules are
bluish-purple with methylene blue, and dark purple by Albert
staining method.
methylene blue stain
Albert stain
metachromatic granules
Culture
•Aerobic and facultative anaerobic, growing well at
37C on blood- or serum-containing medium.
•Loeffler’s serum slant: not a selective medium but
gives abundant growth and typical morphology of the
bacillus.
•Blood tellurite agar: a selective medium because
tellurite can suppress the growth of normal flora in
throat, and the colonies are black-colored.
• Pediatric
• Symptoms:
formation of a false membrane in the throat
high fever
difficulty in breathing
Pseudomembrane
• Diphtheria is a typical toxigenic infectious disease.
Diphtheria toxin is the major virulent factor.
•Effective antitoxin therapy and successful toxoid
vaccine available.
• A rare disease,
populations.
• High death rate.
still
occur
in
non-immunized
Virulent factor: diphtheria toxin
The toxin encoding gene (tox) is carried byβcorynebacteriophage
only the bacillus infected by the phage and
committed lysogenic conversion produce diphtheria
toxin
The regulation of tox gene expression is mediated by
an iron-activated inhibitor (DtxR) which is
chromosomally encoded by C. diphtheria
EM of ß-corynebacteriophage carrying tox gene
◆ many exotoxins are called as A-B type toxins because they
consist of A and B subunits. The B subunit generally mediates
the toxin complex molecule to adhere and then enter the host
cell. A subunit provides the toxic activity.
Cell surface
Toxic
Binding
A
B
Diphtheria toxin is an A-B type toxin
A diphtheria toxin molecule contains
535 amino acids with subunit A and
subunit B.
Subunit B contains receptor binding and
transmembrane domains.
Subunit A contains catalytic domain. It
inactivate elongation factor 2 (EF2). Thus
the protein synthesis is suppressed.
The toxin is very toxic to humans and
many animals (e.g.,monkey, rabbit,
guinea-pig).
Particularly harmful to heart and central
nervous system.
The lethal dose is as low as 100 to 150
ng/kg.
Clinical Manifestations
• Corynebacterium diphtheriae can cause pharyngitis
which leads to formation of thick grey membrane
called pseudo-membrane. The pseudo-membrane is
composed of fibrin, dead epithelial cells, bacteria and
neutrophils. It exfoliates easily and then blocks the
airway which results in hypoxia (缺氧) and suffocation
(窒息).
• As a result of the action of diphtheria toxin to
peripheral motor neurons and myocardium, lifethreatening systemic complications (principally loss of
motor function and congestive heart failure) may
develop.
Diagnosis of diphtheria requires
laboratory confirmation of toxigenic
Corynebacterium diphtheriae.
It is usually determined by in vitro tests
such as Elek immunodiffusion test.
Immunoprecipitation line
Diphtheria patients must be promptly treated with
antitoxin to neutralize circulating (free) diphtheria
toxin. Antitoxin cannot neutralize the toxicity of
exotoxin that has bound to the host cells.
Antibiotics (penicillin and erythromycin) are used as
part of the treatment.
Diphtheria toxoid is a component of the DPT vaccine,
which includes diphtheria toxoid, killed whole cell
pertussis（Bordetella pertussis） and tetanus toxoid
（Clostridium tetani ）.
Mycobacteria
Acid-fast bacilli
Mycobacteria do not stain readily.
However, if these bacteria are once
stained by Ziel-Neelsen Acid-Fast Stain
method, they are able to resist
decolorization by acid or alcohol.
Among the species of Mycobacteria, M.
tuberculosis is the most important pathogen
that causes human tuberculosis.
M. bovis is a zoonotic causative agent of
cattle tuberculosis. It also causes human
tuberculosis.
M. leprae is the causative agent of leprosy
which is still present in the third world.
Morphology and culture
• Mycobacterium tuberculosis has the typical
morphology and acid-fast property of Mycobacteria.
•Lowenstein-Jensen medium is the widely used
medium for cultivating Mycobacterium tuberculosis.
In this medium, malachite green (孔雀绿) is included
to inhibit other bacteria.
•The growth rate is slower than most other bacteria.
The doubling time is about 18 hours.
The colonies are formed after 3-6 weeks of incubation on
Lowenstein-Jensen medium. The colonies are buff colored
and dry breadcrumb-like.
Variation
Drug resistance variation: Many clinical
isolates of Mycobacterium tuberculosis
are drug-resistant.
Virulence variation: BCG (Bacillus
Calmette-Guerin) is the human
tuberculosis vaccine. It is a preparation
of live attenuated Mycobacterium bovis.
L-form: a form that lacks the cell wall
Virulent factors
Produce no endotoxin, exotoxin or invasive
enzymes
The pathogenicity is due to cell wall
components, and substances inducing
immunopathological reactions.
Virulent factors
Lipids in the cell wall
Cord factor: lipoarabinomannan, damages
mitochondria
Phosphatide: causes the formation of
tubercule and granuloma
Sulfatides: inhibits phagosome-lysosome
fusion
Wax D: serves as an “adjuvant” that
induce hypersensitive reaction.
Virulent factors
Proteins
e.g., Tuberculin mixed with wax D can
induce hypersensitive reaction.
Polysaccharide
Bound to wax D to induce the infiltration
(浸润) of inflammatory cells.
Pathogenesis
Mycobacterium tuberculosis enters alveoli by airborne transmission
It resists destruction by alveolar macrophages and propagate
Tissue destruction
results from cellmediated
hypersensitivity.
To cause primary tuberculosis
To spread to lymph nodes
To enter bloodstream
and seed other organs
To reseed the
lungs
To lead to post-primary tuberculosis
Cell-mediated
immune response
stops cycle of
destruction and
spread.
Transmission and infection
• Mycobacterium tuberculosis is transmitted through
droplet and airborne dust containing the microbe.
• Mycobacterium tuberculosis has multiple routes
invading human body, such as respiratory tract,
intestinal
tract
and
skin,
to
cause
various
tuberculosis.
• Among
the
different
tuberculosis,
tuberculosis is the most common form.
pulmonary
• Bacteriological
examination:
the
definitive
diagnosis for tuberculosis based on the presence of
acid-fast
bacteria
in
the
sputum
(pulmonary
tuberculosis) and in other clinical samples.
• Tuberculin skin test: is a method of identifying
persons who once infected with Mycobacterium
tuberculosis.
• Tuberculin skin test is performed by an intradermal
injection of 5 tuberculin units / 0.1 ml of purified protein
derivative (PPD) into forearm. The result (diameter of
induration around the site of injection) is read 48 to 72
hours after the injection.
•A tuberculin reaction of >= 5 mm of induration is
classified as positive result. However, a positive test
only means to exposure once to the microorganism but
does not certainly indicate active disease.
• Prevention: Vaccination of BCG vaccine.
• Treatment: Antibiotics used to treat tuberculosis
such as streptomycin, isoniazid, rifampin etc., are
quite different from those used to treat other
bacterial infectious diseases. Clinically, several antituberculosis drugs must be simultaneously applied
to avoid drug resistance and to increase efficiency
of therapy.