Lab Eleven:-
________________
Medical Microbiology
Prepared by: Luma J. Witwit
Mycobacteria
Tubercle bacilli are slender, straight or slightly curved rods with rounded
end. In sputum and other clinical specimens they may occur singly or in
small clumps. True branching is occasionally seen in old cultures and in
smears from caseous lymph nodes. They are non- motile, non- sporing,
non- capsulated and acid fast. The Ziehl- Neelsen acid fast stain is useful
in staining organisms from cultures or from clinical material. With this
stain, the tubercle bacilli stain bright red, While the tissue cells and other
organisms are stained blue. Tubercle bacilli are gram-positive but it is
difficult to stain them with gram stain. This is because of the failure of
the dye to penetrate the cell wall.
Most Mycobacteria are found inhabitats such as water or soil. However,
few are intracellular pathogens of animals and humans.
Mycobacterium tuberculosis, along with M.bovis, M.africanum, and
M.microti all cause the disease known as tuberculosis (TB) and are
members of the tuberculosis species complex.
Each member of the TB complex is pathogenic but M. tuberculosis is
pathogenic for humans while M. bovis is usually pathogenic for animals.
Tuberculosis complex organisms are:
1- Obligate aerobes growing most successfully in tissues with a high
oxygen content, such as the lungs.
2- Facultative intracellular pathogens usually infecting mono nuclear
phagocytes (e.g macrophages).
3- Slow-growing with a generation time of 12 to 18 hours.
4- Hydrophobic with a high lipid content in the cell wall. They are
impermeable to the usual stains e.g. Grams stain.
5- Known acid fast bacilli because of their lipid-rich cell walls, which
are relatively impermeable to various basic dyes unless the dyes are
combined with phenol. The cells resist decolorization with acidified
organic solvents; therefore, are called "acid-fast".
-1-
Lab Eleven:-
________________
Medical Microbiology
Prepared by: Luma J. Witwit
Diagnosis of Tuberculosis:
Diagnosis of tuberculosis is made by a positive tuberculin skin test, an
immuno reaction to a small quantity of tuberculosis antigens. It can be
confirmed by (X rays) of the chest and microscopic examination of
sputum.
Detection of significant numbers of acid-fast bacilli using (Ziehl-Neelsen
stain ) in sputum or tissue samples is considered a positive diagnosis,
laboratory culture of the bacterium (difficult, dangerous and slow-takes at
least 4 weeks).
Direct microscopic examination:
Sputum : a film is prepared from the purulent portion of the sputum and
stained by the Ziehl-Neelsen method.
Cultivation:
Collect sputum not saliva, liquefy the sputum by (1%) of (puffered
pancreatin ), mix the sputum with (trisodium phosphate ) to kill other
bacteria, the sediment used to inoculate the (Lowenstein-Jensen slant ) or
(serum slopes) finally detect the growth checking weekly. LJ medium
may made selective by addition of some antibiotics in addition to
malachite green to suppress bacterial and fungal contaminants. M.
tuberculosis form rough, tough and buff colonies on this media.
Injection in Guinea pig :
The Guinea pig is highly susceptible to infection by both M. tuberculosis
and M. bovis . after subcutaneous injection of an infected specimens, a
locale swelling appears with in a few days , which proceeds to caseate
and finally ulcerate . the organism spreads to the lymph nodes , spleen ,
liver and peritoneum . the animal dies with in 6-15 weeks.
Because, modern methods of culture are so efficient, the inoculation of
sputum into animals is rarely indicated.
-2-
Lab Eleven:-
________________
Medical Microbiology
Prepared by: Luma J. Witwit
Examination of other specimens:1- Fluids (pleural, pericardial, synovial, peritonial fluids.
2- Bronchial washing liquids.
3- Gastric lavage.
4- Urine.
These are centrifuged and stained the deposit by Ziehl- Neelsen method.
5- Tissue biopsies.
6- Pus and bone marrow.
Are stained directly by usual way.
Leprosy:
Is caused by the organism Mycobacterium leprae. Children are more
susceptible than adults to contracting the disease.
Leprosy has two common forms, tuberculoid and Lepromatous. Both
forms produce lesions on the skin, but the Lepromatous form is most
sever, producing large disfiguring nodules.
All forms of the disease eventually cause peripheral neurological
damage (nerve damage in the extremities by sensory loss in the skin and
muscle weakness).
People with long-term leprosy may lose the use their hands or feet due to
repeated injury resulting from lack of sensation.
Lepromin skin test :can be used to distinguish lepromatous from
tuberculoid leprosy.
Primery growths may be obtained on serum media or on medium
containing egg yolk.
In secondary culture, growths may result on ordinary media (agar, broth)
with 5-6 percent glycerol added.
-3-