Download Mycobacterium tuberculosis

Bacteriology
lecture no.3
Mycobacterium
*General characteristics :
1. Species belonging to the genus Mycobacterium are very thin , rod shaped , and non – motile , they are obligate aerobes .
2. Mycobacterium spp . have unusual cell wall structure that contains N –
glycolylmuramic acid and has a very high lipid content .
3. Because of this cell wall structure , Mycobacteria are difficult to stain
with commonly used basic aniline dyes , such as these used in the Gram
stain .
4. These organisms resist decolorization
by acidified alcohol (3%
hydrochloric acid ) after prolonged application of a basic fuchsin dye
or with heating of this dye following its application .
5. This important property of mycobacteria that is dependent on its cell
wall is referred to as acid – fastness , and this feature can be
distinguished it from all other species .
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6. Another important feature of these organisms is that they grow more
slowly than most other human pathogenic bacteria because of their
hydrophobic cell surface , because of this hydrophobicity , organisms
tend to clump , so that nutrients are not easily allowed into the cell .
7. Growth are slow or very slow , with colonies becoming visible in 2 to 60
days at optimum temperature .
8. Currently , there are more than 100 recognized or proposed species in
the genus Mycobacterium .
9. There species produce a spectrum of infections in humans and animals
ranging from localized lesions to disseminated disease .
10. Some species cause only human disease , others have been isolated
from a wide variety of animals , many species are also found in water
and soil .
*Classification of Mycobacterium
For the most part , Mycobacteria can be divided into two major
groups based on fundamental differences in epidemiology and
association .with disease
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* Notes: Species always considered pathogens
1. Mycobacterium tuberculosis in humans .
2. Mycobacterium leprae in humans .
3. Mycobacterium bovis in human and cattle .
4- Mycobacterium ovium complex considered as potential pathogen in
human and it present in soil , water , birds , fowl , swine , cattle and
cause disease in immunocompromised patient .
* Mycobacterium tuberculosis
A- Typical organisms .
1. Tubercle bacilli are thin straight rods .
2. Mycobacteria cannot be classified as either gram- positive or gramnegative .
3. Once stained by basic dyes they cannot be decolorized by alcohol ,
regardless of treatment with iodine .
4. True tubercle bacilli are characterized by acid-fastness, i.e., 95%ethyl
alcohol containing 3% hydrochloric acid ( acid – alcohol ) quickly
decolorizes all bacteria except the Mycobacteria .
5. The ziehl- Neelsen technique of staining is employed for identification
of acid – fast bacteria .
6. Structure of cell wall of Mycobacterium essentially put it into grampositive bacteria ,but it cannot take the gram- stain .
B- Growth characteristics .
1. Mycobacteria are obligate aerobes and derive , energy from the
oxidation of many simple carbon compounds .
2. Biochemical activities are not characteristics .
3. The growth rate is much slower than that of most bacteria .
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C-Reaction to physical and chemical agents. .
1. Mycobacteria tend to be more resistant to chemical agents than
other bacteria because of the hydrophobic nature of the cell
surface and their clumped growth .
2. Dyes e.g. . malachite green or antibacterial agents e.g.pencillin
that are bacteriostatic to other bacteria can be incorporated into
media without inhibiting the growth of tubercle bacilli .
3. Acids and alkalies permit the survival of some exposed bacilli
and are used to help eliminate contamination organisms and for
concentration of clinical specimens .
4. Tubercle bacilli are resistant to drying and survive for long
periods in dried sputum .
*Constituents of Tubercle Bacilli .
The constituents that are listed below found mainly in cell walls ,
these constituent can induce delayed hypersensitivity , and some
resistant to infection .
a- Lipids
Mycobacteria are rich in lipids ,these includes mycolic
acids (long _chain fatty acid C78-C90),waxes and phosphatides. IN
the cell, the lipids
are largely bound to protein and
polysaccharides.
-muramyl dipeptide (from peptidoglycan)complexed with mycolic
acids can cause granuloma formation.
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-This
structure is responsible for acid fastness of mycobacterium
and also responsible for its virulence.
b-protien:Each type of
mycobacterium contains several proteins
that elicit the tuberculin reaction
c-Polysaccharides
mycobacteria contain a variety of a
polysaccharide their role in pathogenesis of disease is uncertain .it
can induce immediate type of hypersensitivity
*pathogenesis
1. mycobacteria are emitted in droplet <25µm in diameter when infected
person cough ,sneeze, or speak.
2. The droplet evaporate leaving organisms which are small enough, when
inhaled, to be deposited in alveoli.
3. Once inside the alveoli ,the hosts immune system responds by release of
cytokines and lymphokines that stimulate monocytes and macrophages.
4. mycobacteria begin to multiply within macrophages.
5. Some of the macrophages develop an enhanced ability to kill the
organism while others may be killed by the bacilli.
6. After 1-2 months following exposure ,pathogenic lesion associated with
infection ,appear in the lung.
7. Resistance and hypersensitivity of the host greatly influence
development of disease and the type of lesions that are seen.
*Pathology.
-The
production and development of lesions and their healing or
progression
are
determined
chiefly
by
1- the number of mycobacterium in the inoculum and their subsequent
multiplication .
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2-The type of host.
-Infection with mycobacterium tuberculosis can produce tow principal lesions.
A-Exudative type. This consist of an acute inflammatory reaction
,with edema fluid ,polymorphonuclear leukocytes, and later
monocytes around the tubercle bacilli.
-this type is seen particularly in the lung tissue ,where it resembles
bacterial pneumonia.
-It may heal by resolution ,or it may lead to massive necrosis of
tissue ,or it may progress into productive type.
B-Productive type.
-when fully developed ,this lesion is called as chronic granuloma.
-chronic granuloma, consists of 3 zones.
1-central area of large ,multinucleated giant cells containing
tubercle bacilli.
2-a mid-zone of pale epitheloid cells ,often arranged radially.
3-a peripheral zone of fibroblasts ,lymphocytes and monocytes.
-later, peripheral fibrous tissue develops ,and the central area
undergoes caseation necrosis. This lesion is called a tubercle.
-it may subsequently heal by fibrosis or calcification .
-Tubercle bacilli spread in the host by direct extension, through the
lymphatic channels and bloodstream ,and via the bronchi and
gastrointestinal tract.
-the bacilli may spread and reach the bloodstream ,which in turn
distributes bacilli to all organs (military distribution)
-If a caseating lesion discharges it’s content into a bronchus ,they
are aspirated and distributed to other parts of the lung or are
swallowed and passed into the stomach or intestine .
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-Mycobacteria , they reside principally intracellularly in monocytes
reticuloendothelial cells , and giant cells , this intracellular location
makes chemotherapy difficult and favors microbial persistence .
- When a host has first contact with tubercle bacilli , the following
features are usually observed .
1. An acute exudative lesion develops and rapidly spreads to the lymph tics
and regional lymph nodes .
2. The lymph nodes undergoes massive caseation , which usually calcifies
(Ghon lesion ) .
3. The tuberculin test becomes positive , all these above is called primary
infection that occur early in childhood .
- In primary infections , the involvement may be in any part of the lung
but it is most often at the base .
- The reactivation type is usually caused by tubercle bacilli that have
survived in the primary lesion .
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- Reactivation tuberculosis is characterized by chronic tissue lesions the
formation of tubercles ,caseation and fibrosis .
- The reactivation type almost always begins at the apex of the lung ,
where the O2 tension is highest .
*Tuberculin test
A. Material .
- Old tuberculin is a concentrated filtrate of broth in which tubercle
bacilli have grown for 6 weeks .
- In addition to the reactive tuberculoproteins , this material contains
a variety of other constituents of tubercle bacilli and of growth
medium .
- A purified protein derivative (PPD) is obtained by chemical
fractionation of old tuberculin .
B. Dose of tuberculin .
- A large amount of tuberculin injected into a hypersensitive
host many give rise to severe local reactions and flare-up of
inflammation and necrosis at the main sites of infection .
- For this reason ,tuberculin test in surveys employ 5Tu while
in persons suspected of extreme hypersensitivity skin testing
is begun with 1 Tu .
- 250Tu is administered only if the reaction to 5Tu is negative .
- The volume is usually 0.1ml injected intracutaneously .
C. Reactions to tuberculin .
- In an individual who has not had contact with mycobacteria , there is
no reaction to PPD .
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- An individual who has had a primary infection with tubercle bacilli
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develops induration , edema , erythema in 24-48 hours ,and ,with very
intense reactions , even central necrosis .
It is considered +ve if the injection of 5Tu is followed by induration
10mm or more in diameter , +ve test persist for several days .
Tuberculin test may be negative in the presence of tubercles infection
in the following conditions .
1- measles . 2- Hodgkin disease .
3- sarcoidosis .
4- AIDS .
5-other immunosuppression conditions .
After BCG vaccination , people convert to a positive test ,but this
may last for only 3-7 years .
So positive result indicated that the patient is either had a past
infection or he is vaccinated and at risk of reactivation .
Negative result indicated that this person has no infection and
because person who live in endemic area or had a vaccination develop
positive result so this test is not good in discovery of infection
specially in an endemic area .
So in effort has developed to improve diagnostic accuracy whole –
blood gamma interferon release assays have been commercially ,
these assays are based on the hosts immune .
Responses to specific M. Tuberculosis antigens ESAT-6 and CFP-10,
which are absent from most nontuberculous Mycobacteria and BCG .
Gamma Interferon Release test detect interferon gamma that is
released by sensitized CD4 T cells in response to these antigens .
*Mycobacterium Leprae
1. This organism was described by Hansen in 1873, 9years before Koch's
discovery of the tubercle bacillus .
2. It has not been cultivated on nonliving bacteriologic media.
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3. It cause leprosy ,there are more than 10 million cases of leprosy
,mainly in Asia.
4. It is typically acid fast bacilli ,usually occurs singly, in parallel bundles
,or in globular masses.
5. Samples is taken from scraping from skin or mucous membrane
particularly from the nasal septum in case of lepromatous leprosy.
6. The bacilli are often found within the endothelial cells of blood vessels
or in mononuclear cells.
7. The lesions involve the cooler tissue of the body ,skin superficial
nerves ,nose,pharynx,larynx,eyes,and testicles.
8. The skin lesions may occur as pale ,anesthetic macular lesions 1-10cm
in diameter, diffuse or discrete erythematous, infiltrated nodules 1-5
cm in diameter or a diffuse skin infiltration.
9. Nerves are always involved with resultant nerve thickness and
neurological disturbances are manifested by anesthesia, neuritis
,paresthesia .with bone resorption and shortening of digit.
10. The disfigurement due to the skin infiltration and nerve involvement
may be extreme.
11. Disease is divided into two major types ,lepromatous and tuberculoid
,with several intermediate stages.
12. In Lepromatous leprosy the course of disease is progressive ,and
malignant with nodular skin ,lesion slow symmetric nerve involvement
and abundant acid –fast bacilli in the skin lesions, continuous
bacteremia and negative lepromin skin test ,and cell mediated
immunity is markedly deficient due to infiltration of skin with T-cell
suppressor cells .
13. Tuberculoid type ,the course is benign and nonprogressive with
macular skin lesions ,severe asymmetrical nerve involvement of
sudden onset with a few bacilli present in the lesions ,and a positive
lepromin skin test.
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14. In tuberculoid leprosy ,cell-mediated immunity is intact and the skin
is infiltrated with helper T-cells
15. Diagnosis is made by scalpel blade from skin or nasal mucosa or from
biopsy of earlobe skin ,smear is put on a slide and stained by the ZiehlNeelsen stain
16. No serological test are of value.
17. Nontreponemal serologic test for syphilis frequently yield false –
positive result in leprosy .
18. Transmission of leprosy is most likely to occur when small children
are exposed for prolonged period to heavy shedders of bacilli
19. Nasal secretion are the most likely infectious material for family
contact .incubation period 2-10 years.
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