LECTURE : MYCOBACTERIUM
Diseases: Tuberculosis &
1. Mycobacterium tuberculosis
2. Mycobacterium tuberculosis:
M. Africanum. M. Asian. M.
bovis. M. Avium.
Prof. Abbas Hayat
• 8000-4000 B.C. M. bovis causing TB in animals.
• 5000-1000 B.C, domestication of cattle, human
infection by M. bovis likely through milk ingestion.
1000 BC, widespread pulmonary TB emerged.
• M.tuberculosis, a specialized form of M.bovis
developed among milk-drinking Indo-Europeans
who then spread the disease during their
migration into Western Europe and Eurasia.
• After 1000 B.C. M.TB causing pulmonary TB had
spread throughout the known world.
• 668-626 BC. The classic TB signs--cough,
expectoration, hemoptysis, wasting of the body,
were well recognized. The earliest written
evidence of pulmonary TB was from the library of
the Assyrian king Assurbanipal (668-626 BC):
• 1600 TB responsible for 20% deaths in London.
• 1800 over 30 % deaths in Paris.
• 1865, French military doctor Jean-Antoine
Villemin transmitted organism from one animal to
• 1882 Robert Koch isolated tubercle bacillus.
• 1890 Koch found tuberculin now used to identify
existence of TB.
• 1905, Koch won Nobel Prize for tuberculin.
• 1943. Streptomycin, purified from
• 1949 Following streptomycin, p-amino salicylic
• 1952 isoniazid 1954 pyrazinamide, 1955
cycloserine, 1962 ethambutol 1963 a rifampicin;
were introduced as anti-TB agents. .
• At time of discovery of bacillus, 1/5 people
developed TB during their lifetime.
1. When a guinea pig is injected subcutaneously
with virulent tubercle bacilli, the puncture wound
heals quickly, but a nodule forms at the site of
injection in two weeks. This nodule ulcerates and
the ulcer does not heal. The regional lymph
nodes develop tubercles and caseates
2. When the same animal is injected with tubercle
bacilli in another part of the body, 6-8 weeks
after there is rapid necrosis of skin and tissue,
but the ulcer heals rapidly and regional lymph
nodes do not become infected.
INTRODUCTION to Tuberculosis.
• Someone infected with tuberculosis every second.
• One third of the world's population infected with
tuberculosis complex bacteria.
• Left untreated, one person with tuberculosis will
infect 10-15 people per year. .
• Great concern: New strains; .WHO reports
death with "multi-drug resistant TB" M.D.R.T.in the
U.S. was approximately70 percent.
• Diagnosis to death: four to sixteen weeks.
• 90 million new tuberculosis cases and 30 million
• It strikes people of all races, ages, and income
levels. Higher risk. HIV infection, Close contacts
with infectious TB;
• Poor; Homeless; Prisons; Alcoholics;
Elderly & Health care workers.
AT PRESENT TB is a global emergency according to
3 million cases in Pakistan.
• MORPHOLOGY IDENTIFICATION &
• Zeihl Nelson Staining
• Acid Fast Bacilli: Retain Carbol Fuschin
stain & are not decolorized by Ethanol –
• Counter stain is Methylene blue, (RED
RODS AGAINST BLUE BACKGROUND)
• High lipid content in cell wall 66 % make
them acid fast and heating required for
• Neither Gram positive nor negative.
Drop suspension onto slide
Air dry slide 10 minutes at 60 °C, heat-fix slide 10 minutes at 90 °C
Flood slide with Carbol Fuchsin
Hold flame beneath the slide until steam appears but do not allow it to boil
Allow hot slide to sit for 3 to 5 minutes, rinse with tap water
Flood slide with 30% hydrochloric acid in isopropol alcohol
Allow to sit 1 minute, rinse with tap water
Flood slide with Methylene Blue
Allow to sit 1 minute, rinse with tap water Blot dry
View under oil immersion lens
B. Auramine Florescence Stain.
Fluorescent microscope and Rhodamin and
• Grow slowly. Doubling time 18 hours;
Cultures require 6-8 weeks.
• CULTURE: Lowenstein –Jenson medium
(L.J) (egg yolk and Malachite green dye).
• Obligate Aerobe: Predilection for upper
lobe lungs, and Kidney.
Contains complex lipids
1. Mycolic acids : Acid fastness
2. Wax D: Freunds adjuvant
3. Phospholipids: Caseation necrosis.
4. Glycolipid : Cord factor.
• Proteins + Waxes: elicit
• Resistant to Acids and
Alkalis and Drying.
"Resistance vs. Susceptibility``
• Tissue destruction results from
• Mycobacteria inflammatory lesion
granulomatous lesion characterized by a
mononuclear cell infiltrate surrounding a
core of degenerating epithelioid and
multinucleated giant (Langhans) cells.
This lesion (called a tubercle) fibroblasts,
center progresses to caseous necrosis.
Liquefaction of caseous material → erosion
of the tubercle → cavitations and the release
of massive numbers of bacilli into the
• In resistant host, the tubercle → calcified.
• Early in infection, Mycobacteria → the lymphatics
to the hilar or mediastinal lymph nodes. →thoracic
duct → blood stream, or directly into the
circulation by erosion of the developing tubercle
into a pulmonary vessel.
• Extra pulmonary hematogenous dissemination. →
(e.g., spleen, liver, and kidneys) and, eventually,
reinoculation of the lungs.
• GHON COMPLEX (Primary lesion + Draining
• GHON LESION (Reinfection or Reactivation
without involvement of draining lymph nodes)
PRIMARY & REACTIVATION DISEASE
• Tissue destruction results from presence of
Organism & Host response (cell-mediated
Two types of lesions:
1. Exudative Lesions: Acute inflammatory response
Primary lesion: lower lobes in lungs: Parenchymal
exudative lesion and the draining lymph nodes
are called GHON COMPLEX.
2. Granulomatous Lesions: Central area of Giant
cells containing tubercle bacilli, surrounded by
Reactivation lesions: In apices, also Kidney Brain &
INGESTION OF UNPASTEURIZED MILK
(M.Bovis) gastrointestinal tract.
• Susceptibility is influenced by genetic and ethnic
factors. Acquired resistance is mediated by T
lymphocytes, which lyses infected macrophages
directly or activate them via soluble mediators
(e.g., gamma interferon) to destroy intracellular
bacilli; antibodies play no protective role.
• Clinical signs and symptoms develop in only a
small proportion (5-10 percent) of infected healthy
• Pulmonary disease; prominent symptoms are
chronic, productive cough, low-grade fever, night
sweats, easy fatigability, and weight loss.
• Extra pulmonary manifestations.
• Lymphadenitis; kidney, bone, or joint involvement;
meningitis; or disseminated (miliary) disease.
``The patient coughs frequently; his sputum is thick
and sometimes contains blood. His breathing is
like a flute. His skin is cold, but his feet are hot.
He sweats greatly and his heart is much
disturbed. When the disease is extremely grave,
he suffers from diarrhea.``
• Tuberculosis primarily affects the lower
respiratory system and is characterized by a
chronic productive cough, low-grade fever,
• Night sweats, and weight loss.
• The Mantoux test: intradermal injection of a
measured volume (0.1 ml) containing a
specified quantity (5 tuberculin units) of
PPD. The transverse diameter of induration
is measured 48 to 72 hours later.
• Interpretation varies, as shown in MANTOUX
• 5 T.U. 25 T.U. 250 T.U intradermal
• 48 to 72 hours later 5-10 mm of induration
Interpretation: Positive Test means a person has been exposed,
Negative means Anergy, non exposure defective Cell Mediated
Immunity or Miliary Tuberculosis.
• Sputum, Bronchial, Gastric washings,
Pleural fluid, urine, cerebrospinal fluid
• Biopsy material: endometrial; lymph nodes,
other tissues etc.
• Stained and cultured for acid-fast
bacilli. Culture and identification of
Mycobacteria in such specimens are
mandatory for diagnosis.
3. Commercial chemiluminescent
DNA probes, gas-liquid
chromatography, and thin-layer
identification of a few species of
mycobacteria within hours after
sufficient growth is present on
solid or in a liquid medium
• Aggressive prophylactic chemotherapy in
• In individuals with clinical disease, short term (6-9
month) ambulatory therapy with so-called first-line
anti-mycobacterial drugs, such as isoniazid,
rifampin, pyrazinamide, and ethambutol, results in
disappearance of viable tubercle bacilli from the
sputum, rendering the patient noninfectious.
• Directly observed therapy (DOT) has been
instituted in high prevalence areas, especially
among non compliant patients, as the only reliable
means of ensuring that patients complete their
• the patient is cured.
• the spread of disease is stopped.
• MDR-TB is prevented.
• DOTS has been tested in New
York, Tanzania, Indonesia, Peru,
and China with good results.
According to a report published
in the March 10 issue of the
Archives of Internal Medicine,
the Program resulted in a 52%
decrease in patients with MDR-
TB in New York between 1991
Development of MDRT
• 1960s, 1-2% of isolates were resistant to 2+
• 1970s, 3-5% of isolates were resistant to 2+
• 1986, no more national drug-resistance
• 1991, 33% of isolates resistant to 1+ drugs,
13% resistant to the 4 front-line drugs.
• When resistance to two or more of the first
line drugs is detected, additional drugs
(ethionamide, streptomycin, ciprofloxacin)
may be added to the regimen.
• Following streptomycin, p-amino
salicylic acid (1949),
(1954), cycloserine (1955),
ethambutol (1962) and
rifampin(rifampicin; 1963) were
introduced as anti-TB agents.
Amino glycosides such
kanamycin and amikacin, and the
newer quinolones (e.g.ofloxacin
and ciprofloxacin) are only used in
drug resistance situations.
• Two properties of anti-TB drugs are
important: antibacterial activity, highest
• Isoniazid Rifampin
• and their capacity to inhibit the
development of resistance, the most
effective drugs being
• Isoniazid Rifampin
• The multiple drug regimen described
earlier is very effective-->90% cure rate-
if taken for 6-8 months.
• ``TEST & SLAUGHTER`` Policy
• 1950 – PPD testing of cattle herds and
slaughtering of animals resulted in
elimination of this disease from United
• Improved host resistance,
• Better housing,
• Prophylaxis with Isoniazid.
• In patients exposed to infectious
patients or recent converters and less
than 45 years old.
• A viable, attenuated strain of M bovis,
called bacilli Calmette-Guérin (BCG),
after the French microbiologists. Used
in 120 countries.
• First developed in 1920`s but strain got
contaminated with pathogenic
mycobacterium, vaccination stopped,
till WW II.
• Requires about 200 passages on potato
Nontuberculous Mycobacteria, earlier known
as Anonymous then Atypical Mycobacteria,
now MYCOBACTERIA OTHER THAN
• A crucial difference between M tuberculosis and
Nontuberculous Mycobacteria is lack of
transmission of the latter from patient to patient.
• No evidence that infections are contagious.
• Organisms exist saprophytically in the soil or
water, occasionally in association with some
infected-animal reservoir (e.g., poultry infected
with M avium). Inhalation or ingestion of viable
Mycobacteria or introduction of bacilli through
skin abrasions initiates the infection.
Pathogenesis is similar to Mycobacteria. There
may be granuloma formation
SIGNS & SYMPTOMS
• Patients exhibit lower respiratory
disease similar to tuberculosis (M
kansasii, M avium-intracellulare),
• (M scrofulaceum), skin and soft tissue
infections (M ulcerans, M marinum), or
disseminated disease in persons
infected with HIV.
CLINICAL PRESENTATION OF MYCOBACTERIA
OTHER THAN TUBERCULOSIS (MOTT)
Group 1 (Photocromogens)
• M. kansasii: resembles tuberculosis
• M. marinium: granulomatous ulcerative lesion ``Swimming
pool granuloma`` treatment by tetracycline effective.
Group II (Scotochromogens)
• M. scorfulaceum: cause scrofula;
graunulomatous cervical adenitis in children.
Surgical excision can cure.
Group III (Nonchromogens)
• M. avium- intracellularae pulmonary disease
indistinguishable from tuberculosis esp. in
immunocompromised and HIV; highly resistant to anti
tuberculous drugs 06 drug combination may be required.
Group IV ( Rapid Growing Mycobacteria)
• M. fortuitum-chelonei complex: rarely cause human disease except in
1. Immunocompromised 2.prosthetic heart valves and hip joints...
Frequently resistant may require multiple drug therapy and surgical
• M.Smegmetis : Non pathogenic but confuses in A.F.B. smears.
Treatment and Control
• Many Nontuberculous Mycobacteria are resistant
to commonly used drugs
• Antibiotic regimens may require several (five or
six) drugs including rifampin, which is quite
effective against M kansasii, or clarithromycin,
which has marked activity against the M avium-
• Surgical resection is occasionally recommended
with or without chemotherapy.
• In treating disseminated infections in AIDS
patients, a regimen of five or six drugs, including
clarithromycin, ethambutol and perhaps
rifampicin, should be considered.
TWO TYPES OF DISEASE
1. LEPROMATOUS LEPROSY:
course progressive and malign
with nodular skin lesions,
symmetric nerve involvement
abundant acid fast bacilli in
bacteremia and a negative
lepromin skin test
• 2. TUBERCULOID LEPROSY:
course is benign and
nonprogressive, with macular skin
lesions , severe asymmetric nerve
involvement of sudden onset and
few bacilli present in lesions, and a
Positive lepromin skin test.
• Delayed hypersensitive is markedly
defective in lepromatous leprosy.
• Insidious onset.
• Involve cooler tissues of body i.e. skin
superficial nerves , nose, pharynx, eyes, and
• Skin lesions as pale anesthetic macular lesions
1-10 cm in dia, diffuse or discreet
erythematous, infiltrated nodules 1-5 cm. or a
diffuse skin infiltration.
• Neurological involvement manifested
by nerve infiltration and thickening,
resultant anesthesia, neuritis,
parasthesia , trophic ulcers, and bone
reabsorption with shortenening of
• Disfiguration may be extreme ``Lioness
• Scraping from skin, nasal mucosa, or from
biopsy of the ear lobe skin are smeared on
slide and stained by ZIEHL NELSON
• Biopsy of skin or thickened nerve gives
characteristic histological picture.
• Sulphones i.e dapsone and Rifampin
suppress growth if given for many
• Sulphone resistance emerging .
• Amithiazone substitute drug.
PREVENTION & CONTROL
• In endemic areas removal of young
children from infected families is
employed with some success.
• Chemotherapy of infected cases good
prophylaxis for the community.
• Chemoprophylaxis of contacts with
• Experimental B.C.G. has been used
with possible benefits.