Tuberculosis

‫تپ دق‬

Prof. Dr. Sehar Afshan Naz

M.Phil., PhD
 Tuberculosis
• Tuberculosis (TB) is an infectious disease
caused by Mycobacterium tuberculosis (MTB)
bacteria and characterized by the formation of
tubercles (round nodules) or granulomas in
the lungs. It may be transmitted to other body
parts such as meninges, bones, kidneys, and
lymph nodes.
• Tuberculosis has been present in humans
since ancient times.

• In 2018, there were more than 10 million

cases of active TB which resulted in 1.5 million
deaths.
• Most TB cases occurred in the regions of South-
East Asia (44%), Africa (24%) and the Western
Pacific (18%), with more than 50% of cases being
diagnosed in eight countries: India (27%), China
(9%), Indonesia (8%), the Philippines (6%),
Pakistan (6%), Nigeria (4%) and Bangladesh (4%).
 Clinically Important Mycobacterium
species
• Mycobacterium tuberculosis causes
tuberculosis in human
• M. leprae: Cause Leprosy in humans
• M. bovis: Causes infections in humans and
cattles
• M.kansasii: Causes infections in humans and
cattles
 Etiology
• The main cause of TB is Mycobacterium
tuberculosis (MTB), discovered by Robert Koch

• Straight or slightly curved thin rod-shaped

bacilli.
• Non-sporing, non-motile
• non-capsulated bacteria
• They are neither Gram +ve nor Gram –ve. If a
Gram stain is performed, MTB either stains very
weakly "Gram-positive" or does not retain dye
due to its cell wall's high lipid and mycolic acid

• A key factor of the M. tuberculosis cell

membrane is that over 60% of it is made out of
lipids. The cell wall of M. tuberculosis is unique
in that it has mycolic acids, cord factor, and
mAPG complex.
• Mycolic acids are long fatty acids found in the
cell wall of M. Tuberculosis.

• Because mycolic acids contribute to the waxy-

like coating on the cell surface, the
mycobacterial cells are essentially impervious
to normal gram staining techniques.

• Hence, acid-fast staining is utilized because it

can penetrate through the dense lipid layer
and stain the bacterial cell
• Ziehl-Neelsen stain is used to dye
M. tuberculosis with a special pink
stain called carbon-fuchsin.
Resultantly, the acid-fast stained
bacteria will appear light pink
• https://youtu.be/pm80zK-v3Ac

• They are called Acid Fast Bacilli

(AFB) or Acid Alcohol fast Bacilli
because they are not decolorized
easily by acids or alcohols. This
property is also because of the
high content of mycolic acid
present in its cell wall.
• They appears in single, in
pairs, or aggregates in long
strands known as serpentine
cords
• The cell wall is rich in lipids
and waxes.
• The high content of mycolic
acid (50 to 60 %) present in
cell wall.
• They are wrapped together
due to the presence of fatty
acids known as cord factors.
• Capable of intracellular growth.
• It divides every 16 to 20 hours, which is
an extremely slow rate compared with
other bacteria, which usually divide in
less than an hour.
• They are resistant to disinfectants,
detergents, common antibiotics, dyes,
stains, osmotic lysis, lethal oxidation,
etc.
• They may be obligate pathogen or
opportunistic pathogens.
• Usually infect the mono-nuclear
phagocytes.
• Can also be infected by the
consumption of unpasteurized milk.
• Can survive for weeks in the dust, on
carpets or clothes, for months in
sputum.
• They are also found in soil and water.
 Growth Characteristics on media
• Slow grower even on enriched media.
• Generation time is 14 hours.
• Commonly used media for growth in Lab is
Lowenstein Jensen Medium (L J medium).
• It is an egg-based medium and growth is quite
slow.
• It takes 6-8 weeks to get visual colonies on this
type of media.
M.Tuberculosis on L J Medium
 Virulence factors of M.tuberculosis
• Mycolic acid
• Lipoarabinomannan (LAM)
• Cord Factor
• Sulfolipids
 Mycolic acid

• Waxy layer that protects the bacteria from

many host factors and also to many antibiotics
including beta-lactamases.

• The high content of mycolic acid (50 to 60 %)

present in cell wall.
 Lipoarabinomannan (LAM)

• LAM is structurally and functionally related

to the O antigenic lipopolysaccharides of
other bacteria and can cause suppression
of T-cell proliferation.
 Cord Factor
• Cord factor is a surface glycolipid
and causes M. tuberculosis to grow
serpentine cords.
• Virulent strains of Mycobacterium
tuberculosis possess the cord
factor factor.
• It is toxic to leukocytes, anti
chemotactic, interferes with
mitochondrial function in and
plays a role in the development of
granulomatous lesions
 Sulfolipids

Sulfolipids prevent phagosome-lysosome fusion

so that the organisms are not exposed to
lysosomal enzymes (is important in intracellular
survival)
• Transmission
• When people with active
pulmonary TB cough,
sneeze, speak, sing, or spit,
they expel infectious
aerosol droplets 0.5 to 5.0
µm in diameter.

• A single sneeze can release

up to 40,000 droplets.
Each one of these droplets
may transmit the disease.
• The infectious dose of tuberculosis is very
small (the inhalation of fewer than 10
bacteria may cause an infection).
 Pathogenesis
• The infectious bacilli gain its entry to the
human host from inhalation as droplet nuclei
from the atmosphere.
• The first line of defense of the host tries to
ward off the bacteria by tracheal and
bronchial epithelium.
• When bacteria survive from the first line of
defense, they enter the lungs.
• Alveolar macrophages ingest these tubercle
bacilli; most of these bacilli are destroyed or
inhibited.
• A small number may multiply intracellularly
and are released when the macrophages die.
• If alive, these bacilli may spread by way of
lymphatic channels or through the
bloodstream to more distant tissues and
organs (including areas of the body in which
TB disease is most likely to develop: regional
lymph nodes, apex of the lung, kidneys, brain,
and bone).
• This process of dissemination primes the
immune system for a systemic response.

• Tuberculosis can be of three types

• 1- Primary Tuberculosis
• 2- latent Tuberculosis
• 3- Secondary Tuberculosis
Primary tuberculosis
• When the bacteria are phagocytosed by alveolar
macrophages in the lung, but survives in the
phagosome, and induces a localized cell-
mediated pro-inflammatory response leading to
granuloma formation also known as TUBERCLE.
• The granuloma protects the bacteria and
generally appear after 3 weeks of primary
infection.
• .
• .
• When fully developed, this lesion, a chronic
granuloma, consists of three zones
• a central area of large, multinucleated giant
cells containing tubercle bacilli;
• a mid-zone of pale epithelioid cells often
arranged radially; and
• a peripheral zone of fibroblasts, lymphocytes,
and monocytes.
• The center of the granuloma contains a
mixture of necrotic tissue and dead
macrophages
• Granuloma formation is usually sufficient to
limit the primary infection.
• The lesions become quiescent (dormant) and
surrounding fibroblasts produce dense scar
tissue, which may become calcified called
ghon focus.
• Programmed cell death (apoptosis) of
bacteria-laden macrophages by cytotoxic T
cells and natural killer (NK) cells contributes to
protective immunity by generating a
metabolic burst that kills tubercle bacilli
• 3. Latent tuberculosis
• In most infected individuals, the primary
infection resolves but some residual tubercle
bacilli enter a poorly understood stage of
latency or dormancy.
• This is mostly observed in
immunocompromised cases eg. HIV or aging.
Secondary Tuberculosis
• Reactivation of dormant foci of tubercle
bacilli or exogenous reinfection leads to
post-primary tuberculosis, which differs in
several respects from the primary disease.
• For unknown reasons, reactivation or
reinfection tuberculosis tends to develop in
the upper lobes of the lungs.
 Clinical manifestation
1. Primary or pulmonary tuberculosis
• Primary tuberculosis typically is considered a disease
of the respiratory tract.
• Common presenting symptoms include:
• low-grade fever
• night sweats
• fatigue
• Anorexia (loss of appetite)
• weight loss
• Myalgia ( pain in muscles)
• .
2. Disseminated tuberculosis
• Organs besides the lungs can become involved
after infection with M. tuberculosis
• These organs include the following:
• Genitourinary tract
• Lymph nodes (cervical lymphadenitis)
• The central nervous system (meningitis)
• Bone and joint (arthritis and osteomyelitis)
• Peritoneum
• Pericardium
• Larynx
• Pleural lining (pleuritis)
 Laboratory diagnosis
Collection of Specimen and processing
• sputum, bronchial washings, or biopsies or
early morning gastric aspirates, Cerebrospinal
Fluid (CSF), urine
• Specimens from sputum and other non sterile
sites should be decontaminated with NaOH
(kills many other bacteria and fungi).
• Specimens from sterile sites, such as
cerebrospinal fluid, do not need the
decontamination procedure
• Direct Microscopy
• Detection of the acid-fast property of
mycobacteria to detect them in sputum
and other clinical material by the Ziehl–
Neelsen (ZN) staining technique.
• Red bacilli are seen against the contrasting
background color.
• Slender, straight, or slightly curved rods
with a barrel or beaded appearance.
• Fluorescence microscopy, based on the
same principle of acid-fastness, is
increasingly used and is much less tiring.
 Primary Isolation of Mycobacterium
tuberculosis

• After decontamination with NaOH, the clinical

specimen is inoculated on Lowenstein Jensen (LJ)
medium and incubated for 2 months
• Specimens such as cerebrospinal fluid and tissue
biopsies, which are unlikely to be contaminated, are
inoculated directly onto culture media.
• Dry, rough, raised, wrinkled, off white to buff-
colored colonies on LJ medium, commonly called as
rough, tough, and buff colonies.
BACTEC 460 TB

• The use of the liquid medium with

radiometric growth detection such as has
simplified the culture method.
 Figure: Colony morphology of Mycobacterium
tuberculosis on Lowenstein Jensen (LJ) agar after 8
weeks of medium
 Biochemical Identification
• Different biochemical tests are used in
identification of M.tuberculosis
 Serodiagnosis
• ELISA
• Latex agglutination Test
 Molecular Techniques for Detection
• Polymerase Chain Reaction (PCR)
 Indirect detection method
• Tuberculin test
• A purified protein derivative (PPD) is obtained
by chemical fractionation of old tuberculin.
• PPD is standardized in terms of its biologic
reactivity as tuberculin units (TU).
• Tuberculin tests in surveys use 5 TU in 0.1 mL
solution; in persons suspected of extreme
hypersensitivity.
• After the tuberculin skin test is placed, the
area is examined for the presence of
induration no later than 72 hours
• The result interpretation is given as:
• 5 mm or larger of induration is considered
positive. For patients, at the highest risk of
developing the active disease (eg, HIV-
infected persons, people who have had
exposure to persons with active tuberculosis)
• Larger than 10 mm is considered positive for
persons with an increased probability of
recent infection. This category might include
individuals such as recent immigrants from
high-prevalence countries, injection drug
users, and health care workers with exposure
to tuberculosis.
• For persons at low risk for tuberculosis, 15
mm or larger of induration is considered a
positive test result.
• Positive test results tend to persist for several
days. Weak reactions may disappear more
rapidly.
 Treatment
• The first line of anti-TB agents
that form the core of treatment
regimens are
• Isoniazid (INH)
• Rifampin (RIF)
• Pyrazinamide (PZA)
• Ethambutol (EMB)
• The current World Health
Organization recommendations
are that all new patients with
tuberculosis, should receive a 6-
month course of therapy.
• ).
Resistance may develop during therapy
(acquired resistance) with poor-quality drugs
or inadequate supervision, or patients may be
infected with resistant strains (initial or
primary resistance
 Prevention
• Human tuberculosis is
preventable:

• by the early detection and

effective therapy of the open
or infectious individuals in a
community
• by lowering the chance of
infection by reducing
overcrowding