Mycobacteria

Tuberculosis
Tuberculosis is necrotizing
bacterial infection with waid
distribution. The lungs are most
common by affected (75%) -
pulmonary tuberculosis.
Extra pulmonary tuberculosis
includes: meningitides,
osteomyelitis, miliary
tuberculosis (multisystem
infection – associated with night
sweats, fever and weight loss),
cervical and mesenteric
lymhadenoparthy, abdominal
According to the World Health
Organization (WHO), nearly 2
billion people—one third of the
world's population—have been
exposed to the tuberculosis
pathogen., 8 million people
become ill with tuberculosis,
and 2 million people die from
the disease worldwide. In
2004, around 14.6 million
people had active TB disease
with 9 million new cases. The
annual incidence rate varies
from 356 per 100,000 in Africa
to 41 per 100,000 in the
M. tuberculosis, then known as
the tubercle bacillus, was first
described on 24 March 1882 by
Robert Koch, who subsequently
received the Nobel Prize in
physiology or medicine for this
discovery in 1905; the bacterium
is also known as Koch's bacillus.
 Mycobacterium can be classified into several major
groups
 M. Tuberculosis -95%
 M. Bovis -3-5% (M. Bovis infection is typically
localized to bone marrow and cervical or
mesenteric lymph nodes).
 M.africanum,-2-3%;
 M. leprae which causes Hansen's disease or
leprosy;
 Nontuberculous mycobacterium (NTM) are all the
other mycobacterium which can cause pulmonary
disease resembling tuberculosis, lymphadenitis,
skin disease, or disseminated disease.
 Clinical infection due to other atypical
mycobacterium is rare.
The genus Mycobacterium
consist of non motile, non spor-
forming, non-capsulated, obligate
aerobic.
Classification is based on culture
characteristics, including
nutritional requirement, rate of
growth, pigmental and
biochemical properties.
Classification is based on culture
characteristics, including nutritional
requirement, rate of growth, pigmental
and biochemical properties.
Mycobacteria cannot be classified as
either G+ or G-, and cannot be
decolorized by alcohol, but quickly
decolorized by 3% hydrochloric acid.
The genome of the H37Rv strain
was published in 1998.
Its size is 4 million base pairs, with
3959 genes.
The genome contains 250 genes
involved in fatty acid metabolism,
with 39 of these involved in the
polypeptides metabolism
generating the waxy coat. Such
large numbers of conserved genes
shows the evolutionary importance
of the waxy coat to pathogen
survival.
M. tuberculosis is an obligate
aerobe (weakly Gram-positive
mycobacterium, hence Ziehl-
Neelsen staining, or acid-fast
staining, is used). While
mycobacterium do not seem to fit
the Gram-positive category from an
empirical standpoint (i.e., they do
not retain the crystal violet stain),
they are classified as acid-fast
Gram-positive bacteria due to their
lack of an outer cell membrane
All Mycobacterium species share a
characteristic cell wall, thicker
than in many other bacteria, which
is hydrophobic, waxy, and rich in
mycolic acids/mycolates. The cell
wall consists of the hydrophobic
mycolate layer and a
peptidoglycan layer held together
by a polysaccharide,
arabinogalactan.
M. tuberculosis grow very
slowly
M. tuberculosis divides every
15-20 hours, which is
extremely slow compared to
other bacteria, which tend to
have division times measured
in minutes (E. coli) can divide
roughly every 20
minutes).Isolation of the slow-
growing organisms can
require 3 to 8 weeks of
incubation.
It is a small bacillus that can
withstand weak disinfectants and
can survive in a dry state for weeks.
Its unusual cell wall, rich in lipids
(e.g., mycolic acid), is likely
responsible for this resistance and is
a key virulence factor
They are unusually resistant to
killing by phagocytes.
They are also higly resistant to
drying, but they are killed during
pasteurization of milk (heating to 60
C for 30 minutes).
 The slow growth results from inability to
transport nutrients rapidly across the wax
layer.
 Slow growth causes delay in diagnosis by
laboratory
 Cultures of clinical material are incubated
for up to 8 weeks.The doubling time of
tubercle ,acilli is about 18 hours.
 On agar, colonies of mycobacterium look
like irregular waxy lumps.
 The pigmented mycobacterium produce
yellow carotinoids.
 Colonies of mycobacterium cannot be
dispersed

colonies of
mycobacterium
Colonies of
mycobacterium
 Cellwall of M.tuberculosis composed of
80% of water, 20 % of dry substances.
 Proteins are presented by 9
tuberculoprotein.
 The most important of them is
tuberculin. The immune response is
aimed against this antigen.
Immunization of sensitized organism by
tuberculin causes the allergic reaction.
Tuberculin
 of the host modify them changing their
antigen structure.
Eextracted and purified preparations of
these protein derivatives are used as a
skin test reagent to measure exposure
 Polysaccharides are presented by
arabinogalactate which ensure the
immunogenicity of mycobacterium.
L lipid s are presented by there groups.
First group – acetonsoluble lipids – can
inter in cell membrane of the host
modify them.
Second – phosphatides are
nonantigenici. Inoculation of phospatides
in laboratory animals caused to
formation of caverna cavity.
Virulence factor
Wax C and Wax D
Wax C – is cord-factor. It consist of
micol acid and thregalose.
It is adhesive factor to macrophages,
antiphagocytic factor at the some
time.
Wax D forms the additional layer and
violate the metabolism of the host
cell.
That is the reason why the
mycobacterium grow slowly.
Wax C form virulrnt strains of tubercle
bacillli
“serpentine cord” in which acid-fast bacilli
are arrenged in parallel chains.
Virulence factor
(conclusion)
Wax C – is cord-factor,
antiphagocytic factor and factor
adhesive to macrophages
Wax D - antiphagocytic factor.
Tuberculin - diffuse the immune
response.
Acetonsolable lipids –which
modify host cell membrane and
diffuse the immune response.
 Epidemiology.
 Infection is transmitted by inhalation of
Mycobacterium tuberculosis in aerosols and dust.
 M. tuberculosis is transmitted from person to person
by respiratory aerosol, initial site of infection is the
lung.
 Air-borne transmission of tuberculosis is efficient
because infected people cough up enormous
numbers of mycobacterium, projecting them into the
environment, where their waxy outer coat allows
survive for long periods of time in air and house dust.
 Human are the natural reservoir of M.
tuberculosis. Most transmission occurs by
aerosols generated by the coughing.
 Initial site of infection is the lung.
 M. bovis is found in cow’s
milk,(unpasteurized) can cause
tuberculosis in human.
 Tubercle bacilli do not produce exotoxin
or endototoxin.
 The severe manifestation of tuberculosis
are linked to host reactions to the
organisms, damage is caused by
uncontrolled, progressive, chronic
inflammation and by organism living with
macrophages.
M.tuberculosis infects macrophages and
spread of the organism within the body
occurs by two mechanism.
1.A tubercle can erode into a bronchus and
spread the organism to other part of the
lungs,
To gastrointestinal tract.
2.It can disseminate via bloodstream to
many internal organs.
Macrophages are the primary cells
infected by M. tuberculosis. Early
in infection, tuberculosis bacilli
replicate essentially unchecked,
while later in infection, the T-
helper response stimulates
macrophages to contain the
proliferation of the bacteria.
 M. tuberculosis enters
macrophages by endocytosis
mediated by several macrophage
receptors: mannose receptors bind
lipoarabinomannan, a glycolipid in
the bacterial cell wall.
M. tuberculosis replicates within the
phagosome by blocking fusion of the
phagosome and lysosome,
mycobacterium block phagolysosome
formation.
 M. tuberculosis has several
mechanisms for blocking
phagolysosome formation, including
inhibition of Ca2+ signals and blocking
recruitment and assembly of the proteins
which mediate phagosome-lysosome
fusion.
Thus the earliest stage of primary
tuberculosis (<3 weeks) in the no
sensitized individual is characterized
by proliferation of bacteria in the
pulmonary alveolar macrophages and
airspaces, with resulting bacteremia
and seeding of multiple sites. Despite
the bacteremia, most patients at this
stage are asymptomatic or have a
mild flulike illness.
About 3 weeks after infection, a TH1
response against M. tuberculosis is
mounted that activates
macrophages to become
bactericidal. TH1 cells are stimulated
by mycobacterium antigens drained
to the lymph node, which are
presented with class II major
histocompatibility proteins by
antigen presenting cells.
Differentiation of TH1 cells depends
on the presence of IL-12, which is
produced by antigen presenting
cells that have encountered the
mycobacterium.
Mature TH1 cells, both in lymph nodes
and in the lung, produce IFN-γ. IFN-γ is
the critical mediator which drives
macrophages to become competent to
contain the M. tuberculosis infection.
IFN-γ stimulates formation of the
phagolysosome in infected
macrophages.
 IFN-γ also stimulates expression of
inducible nitric oxide syntheses (iNOS),
which produces nitric oxide (NO). NO
generates reactive nitrogen
intermediates and other free radicals
capable of oxidative destruction of
several mycobacterium constituents,
from cell wall to DNA.
In addition to stimulating macrophages to
kill mycobacterium, the TH1 response
orchestrates the formation of granulomas
and caseous necrosis. Activated
macrophages, stimulated by IFN-γ, produce
TNF, which recruits monocytes. These
monocytes differentiate into the "epithelioid
histiocytes" that characterize the
granulomatous response. In many people,
this response contains the bacteria and
doesn't cause significant tissue destruction
or illness. In other people, the infection
progresses due to age or
immunosuppression, and the ongoing
immune response results in tissue
destruction due to caseation and cavitation.
Granuloma
Гигантские клетки
The combination of a single lesion in
the lung and caseation in the bronchial
lymph nodes is called the Ghon
complex.
Primary tuberculosis –is the disease of
person who are infected for the first
time.
Primary tuberculosis may take two
courses.
In people who are otherwise healthy,
the lesions become fibrotic or calcified.
These lesion usually persist for a life
time and can be seen years later in
chest x-rays.
Ghon complex.
Caverna
In immunocompromised person, the
organisms may invade the bloodstream
and cause disease in almost any organ
of the body. This can lead to a
potentially fatal generalized infection
-disseminated miliary tuberculosis.

Tubercles are visible in many organs,

including the liver, spleen, kidneys,
brain and meninges.
Miliary tuberculosis.
The name “miliary” is derived from
the millet.
The importance of TNF in this
response is underscored by the fact
that patients with rheumatoid
arthritis who are treated with a TNF
antagonist have an increased risk of
tuberculosis reactivation.
In summary, immunity to M.
tuberculosis is primarily mediated
by TH1 cells(T-lymphocytes), which
stimulate macrophages to kill the
bacteria. This immune response,
while largely effective, comes at the
cost of hypersensitivity and the
accompanying tissue destruction
Reactivation of the infection or re-
exposure to the bacilli in a
previously sensitized host results in
rapid mobilization of a defensive
reaction but also increased tissue
necrosis. Just as hypersensitivity
and resistance appear in parallel,
so, too, the loss of hypersensitivity
(indicated by tuberculin negativity
in a previously tuberculin-positive
individual) may be an ominous sign
that resistance to the organism has
faded.
Two substances produced by
macrophages, interleukin -1 and
tumor necrosis factor are know to
contribute to the symptoms of the
disease.
Interleukin -1 acts as the mediator
of the fever experienced by
patients with tuberculosis.
Tumor necrosis factor or cachectin
interferes with lipid metabolism
and lead to severe weight loss.
Humoral immunity does not play
a major role in the immune
response to tuberculosis.
Antibodies appear in the
circulation but do not seem to
play an effective defensive role,
they useful as a diagnostic tool.
Complications of Mycobacterium
tuberculosis infection arise from local
spread or dissemination Body
The organism may disseminate
via the lymphatic's and
bloodstream to other parts of the
body. This usually occurs at the
time of primary infection, and in
this way chronic foci are
established, which may proceed
to necrosis and destruction in, for
example, the kidney.
Alternatively, spread may be by
extension to a lung, into a
bronchus, or into the pleural
 Clinical fingins
 Gastrointestinal TB – is characterized by
abdominal pain and diarrhea accompanied by
more generalized symptos of fever and weight
loss.
 Intestinal obstruction or hemorrhage may
occur.
 The ileocecal region is the site most often
involved.
 Tuberculosis of GI can be caused by either
M.tuberculosis when it is swallowed after
coughed from a lung lesion or by M.bovis when
it is ingested in unpasteurized milk products.
 In renal tuberculosis –dysuria, hematuria,”flank
pain”
 “Sterile pyuria”. The urine contains white blood
diagnosis of tuberculosis
A diagnosis of tuberculosis is suggested
by the clinical signs and symptoms
referred to above, supported by
characteristic changes on chest
radiography and positive skin test
reactivity in the tuberculin (Mantoux)
test.
 Microscopic examination of a smear of
sputum stained by Ziehl-Nielsen's
method or by auramine often reveals
acid-fast rods.
 This result can be obtained within 1 h of
receipt of the specimen in the laboratory.
This is important because Mycobacterium
Sputum stained by Ziehl-
Nielsen's method
Sputum stained by Ziehl-
Nielsen's method
Sputum stained by auramine
Sputum stained by
auramine
diagnosis of tuberculosis
Rapidnon-culture tests to detect
mycobacteria, e.g. using the
polymerase chain reaction (PCR).
Skin test.
Tuberculin skin test is most widely used to
diagnose tuberculosis.
It only detects delayed hypersensitivity,
and does not indicate the presence of
active disease.
The tuberculin skin test is perfomed by
injecting a small amount of PPD, a mixture
of proteins from tubercle bacilli.
A positive test is indicated by reddening
and thickening (indurations) of skin 48-72
hours after injection.
This is due to infiltration of the area by
mononuclear phagocytes and T-cells.
Skin test.
This delayed-type
hypersensitiiivity reaction reflects
the local evens that take place in
the infected tissue.
Specific antituberculous
drugs
Mycobacterium are innately
resistant to most antibacterial
agents, and specific antituberculous
drugs have to be used; these are
reviewed in. combination therapy -
usually three drugs such as
isoniazid, rifampicin, ethambutol to
prevent emergence of resistance
prolonged therapy - minimum 6
months period which is necessary
to eradicate these slow-growing
intracellular organisms.
Tuberculosis is prevented by
improved social conditions,
immunization and
chemoprophylaxis
Immunization with a live attenuated
BCG (bacilli Calmette-Guérin)
vaccine, has been used effectively
in situations where tuberculosis is
prevalent. Immunization, which
confers positive skin test reactivity,
does not prevent infection, but it
allows the body to react quickly to
limit proliferation of the organisms.
In areas where there is a low
prevalence of disease,
immunization has been largely
replaced by chemoprophylaxis.
Prophylaxis with isoniazid for 1
year is recommended for people
who have had close contact with a
case of tuberculosis. It is also
advocated for individuals who
show recent conversion to skin
test positivity, when it is
essentially early treatment of
subclinical infection rather than
prophylaxis.
“Atypical” mycobacteria
 Atypical” mycobacteria are classified into four groups
according to their rate of growth and whether they
produce pigment under certain condition.
 Group I organism (Photochromogens) produce
 a yellow-orange-pigmented colony only when exposed to light.
 M.kansasii causes lung disease clinically resembling
tuberculosis.
 M.marinum causes “swimming pool granuloma”. These
granuloma, ulcerating lesions occur in the skin at the site of
abrasion incurred at swimming pools and aquriums. The
natural habitat of the organism is both fresh and salt water.
 Treatment with tetracycline.
“swimming pool granuloma”.
Atypical mycobacteria
 Group II organism(Scotochromogenes) produce
the pigment chiefly in dark.
 M.scrofulaceum causes scrofula, a
granulomatous cervical adenitis in children. The
organism enters through the oropharyx and infects
the draing lymph nodes.
 Its natural habitat is environmental water souces,
but it has also been isolated as a saprophyte from
the human respiratory tract.
Atypical mycobacteria
 Group III organism(Nonchromogenes)
 M.avium-intracellulare complex (MAI,MAC).
 Is composed of two species, M.avium and
M.intracellulare.
 They cause pulmonare disease clinically
indistinuishable from tuberculosis, primarily in
immunocompromised patients.
 The organisns are widespread in the environment,
including water and soil.
 They are highly resistant to antituberculosis drugs.
Atypical mycobacteria
 Group IV organism (Rapidly Growing
Mycobacteria).
 M. fortuitum, saprophytes, found in soil and water,
rarely cause human disease.
 Infection occure in two population:
 1.Immunocompromised patients
 2.individuals with with indwellig catheters.
 They are often resistant to antituberculosis therapy.