Mycobacterium

tuberculosis

HISTORY

INTRODUCTION

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.

INTRODUCTION

A closely related pathogen,

Mycobacterium bovis, infects a large
number of animal species (chiefly cattle
[Bovidae] and deer [Cervidae]). Bison,
camel, llama, giraffe, kudu, pigs and
badgers can all be infected.
This bacterium also occasionally infects
humans, but this kind of zoonotic
transmission is rare today.
Tuberculosis should not be regarded as a
zoonosis.

EPIDEMIOLOGY

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
Americas
Human are the only reservoir
Person-to-person spread by aerosol

POPULATIONS AT
GREATEST RISK

Homeless persons
Intravenous drug/alcohol abusers
Prison inmates (Russia and other
previous states of the Soviet
Union)
Recent immigrants to the United
States (Asia, Latin America)
HIV-1 infection/AIDS

CLASSIFICATIONS

Mycobacteria cannot be classified

as either G+ or G-,
and cannot be decolorized by
alcohol,
But quickly decolorized by 3%
hydrochloric acid.

CLASSIFICATIONS

Mycobacterium can be classified into

several major groups
M. Tuberculosis -95%
M. Bovis -3-5%
M.africanum -2-3%;
M. leprae ;
Nontuberculous mycobacterium (NTM)
are all the other mycobacterium which
can cause pulmonary disease
resembling tuberculosis, lymphadenitis,
skin disease, or disseminated disease

CLASSIFICATIONS

At present there are more than 70

recognised species of mycobacteria
Usually the non-tubercular are divided
into two large groups: the fast-growing
and the slow-growing.
This name refers to the speed of
formation of visible colonies on solid
culture bases.
Fast growers produce colonies within 7
days, slow growers in more than 7 days.

CLASSIFICATIONS

The slow growing bacteria are divided

further into 3 groups depending on
pigmentation.
Runyon group I contains the
photochromogens, group II contains the
scotochromogens and group III the nonchromogens. Group IV of the nontuberculous mycobacteria contains the
fast growers.
Scotochromogen species produce yelloworange pigment (carotinoids) in the dark.
Photochromogen species produce their

CLASSIFICATIONS

Some bacteria are very closely related

to each other: e.g. Mycobacterium
tuberculosis, M. bovis, M. africanum and
M. microti (the M. tuberculosis
complex); Mycobacterium avium and M.
intracellulare (MAC).
Some mycobacteria cannot as yet be
cultured in vitro (M. leprae).
Some species cause infection in humans
very rarely, but the course of these
infections can be severe (e.g.
Mycobacterium asiaticum).

Pathogenic Mycobacterium spp.

BCG

AIDS
patients

CHARACTERISTICS

M. tuberculosis is an obligate
aerobe (weakly Gram-positive
mycobacterium, hence ZiehlNeelsen 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-

CHARACTERISTICS

Member of the Mycobacterium

tuberculosis complex which includes M.
tuberculosis, M. bovis, M. bovis BCG, M.
africanum, M. ulcerans
An obligate aerobe
Cell wall lipid rich (25-60% dry weight of
the organism)
Virulence factors --- Cord factor, sulfatides

CHARACTERISTICS

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

Lipid-Rich Cell Wall of Mycobacterium

Mycolic acids

CMN Group:
Unusual cell wall
lipids (mycolic
acids,etc.)
(Purified Protein Derivative)

CHARACTERISTICS

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

CHARACTERISTICS

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

CHARACTERISTICS

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 bacilli is
about 18 hours

CHARACTERISTICS

On agar, colonies of
mycobacterium look like irregular
waxy lumps.
The pigmented mycobacterium
produce yellow carotinoids.
Colonies of mycobacterium cannot
be dispersed in a drop of water

Pathogenesis of
Tuberculosis

Inhalation of small (1-5 m) droplet

nuclei containing M. tuberculosis
expelled by coughing, sneezing, or
talking of another individual with
cavitary tuberculosis
Primary infection by M. tuberculosis
of non-immune alveolar
macrophages with unrestrained
proliferation within the infected

Pathogenesis of
Tuberculosis

Dissemination of infected
macrophages through the draining
lymphatics into the circulation
Development within 3-8 weeks of a
CD4+ T cell dependent cellmediated immune response with
granuloma formation and
macrophage activation at sites of
infection

Pathogenesis of
Tuberculosis

Active infection usually

transformed into latent infection
(exceptions: infants, AIDS)
With decrement in T-cell
dependent cell mediated immunity
(years later) infection reactivated
with development of tuberculosis
(HIV infection, diabetes mellitus,
renal disease, cancer, advanced

Clinical Features of
Tuberculosis

Cough one of the earliest signs with

production of sputum as tissue necrosis
progresses
Dyspnea a later symptom indicating
extensive involvement of pulmonary
parenchyma
Fever and weight loss reflecting
systemic actions of IL-1 and TNF-
(cachectin) secreted by activated
macrophages

LABORATORY INVESTIGATION
FOR MYCOBACTERIOSIS
MICROSCOPIC
DIRECT
CONCENTRATED
CULTUR & DST
ANTIBODY DETECTION
CYTOKINE MEASUREMENT
MOLECULAR
PCR
LCR
HIBRIDIZATION

SPECIMEN
Process ASAP, some is contaminated by normal flora
Appropiate volume
Morning sputum and urine is better
Avoid swab
Type of specimen depend upon infection site
Educate patient on proper method of expectoration

SPUTUM SPECIMEN
( NATIONAL CONTROL PROGRAM )
1. 3 Specimens in 2 visits, at time of visit,next morning and at
time of revisit. Intermittent bacterial shedding
2. 3-5 ml in screw cap,wide mouth container
3. Collected at open air or special room
4. Induction : a. Glyceril guaicolate 200 mg at night
b. Mild exercise follow by deep inspiration
c. Physiological saline inhalation
Prepare smear from purulent, caseous or necrotic portion
Add equal volume of 5 % sodium hypochlorite prior to smear preparation, unless for culture

MICROSCOPY

Diagnosis confirmation
Treatment evaluation
Do not differentiate living and dead
bacteria
All mycobacteria,nocardia and
rhodococcus are AFB
Cut off value 5.000-10.000 bacteria
per ml
ZIEHL NEELSEN (HOT)
MODIFIED-ZIEHL NEELSEN ( HOT )
KINYOUN ( COLD )
KINYOUN-GABBETT ( TAN THIAM HOK ) ( COLD )
AURAMINE FLUOROCHROME ( COLD )

Acid-Fast Organisms
Primary

stain binds cell wall

mycolic acids
Intense decolorization does not
release primary stain from the cell
wall of AFB
Color of AFB-based on primary
stain
Counterstain provides contrasting
background

Recording & Reporting by

WHO/IUATLD recommended
method
Acid Fast Bacilli (AFB) appear red or pink colored rod and
the background is stained in blue colour
More than 10 AFB per field in 20 fields
1 10 AFB per field in 50 fields
10 99 AFB per 100 fields
1 9 AFB per 100 fields

( +++ )
( ++ )
(+)

Exact number
Request

repeat specimen
No AFB found in at least 100 fields

(-)

(Never report as M. tuberculosis by smear result only)

LABORATORY
DIAGNOSIS
Acid-fast smear and culture of a
sediment obtained by NaOH
decontamination and centrifugal
concentration of 3-5 early morning
sputum specimens
Acid-fast stains Carbolfuchsin
(Kinyoun), auramine-rhodamine
Polymerase chain reaction (PCR) of firsttime smear positive respiratory
Dr. John
R. Warren, Department of Pathology, Northwestern University
specimens

Feinberg School of Medicine, June 2007

LABORATORY
DIAGNOSIS
Culture of acid-fast bacilli
Egg based medium (LowensteinJensen)
Agar and broth based medium
(Middlebrook)

Dr. John R. Warren, Department of Pathology, Northwestern University

Feinberg School of Medicine, June 2007

Sputum stained by ZiehlNielsen's method

Sputum stained by auramine

Lowenstein-Jensen Egg
Base Medium

Coagulated whole eggs

Potato flour
Glycerol
Defined salts
Malachite Green (0.025 g/100 mL)
(Petragnani 0.052 g/100 mL)
(ATS 0.020 g/100 mL)

Dr. John R. Warren, Department of Pathology, Northwestern University

Feinberg School of Medicine, June 2007

Dr. John R. Warren, Department of Pathology, Northwestern University

Feinberg School of Medicine, June 2007

colonies of
mycobacterium

Colonies of mycobacterium

MEDIA FOR TB CULTURE

Semisyntethic Agar Media (eg
Middlebrook 7H10, 7H11) : contain
defined salts, vitamins, cofactor,
oleic acid, albumin, catalase,
glycerol
Inspissated Egg Media (eg
Lowenstein Jensen) contain defined
salts, complex organic substances
Broth Media (middlebrook 7H9,
Jawetz, Melnick & Adelberg's medical microbiology, Volume 19
By George F.7H12)
Brooks, Karen C. Carroll, Janet S. Butel, Stephen A. Morse

Biochemical Tests for the

Identification of
Mycobacteria

Niacin accumulation
Nitrate reduction
Pyrizinamidase
Tween 80 hydrolysis
Urease
Arylsulfatase
Iron uptake

Differential Characteristics of
Commonly Isolated
Mycobacterium spp.

Mycobacteria Other
Than Tuberculosis
(MOTT)

Infections acquired from an

environmental reservoir of organisms
Human pathogenicity varies for
different species
Recovery in culture indicates
presence of contaminants,
saprophytes, or pathogens
(correlation with clinical presentation
critical)

Runyon Classification of
MOTT Organisms

Photochromogens
Scotochromogens
Nonchromogens
Rapid Growers

Mycobacteria Other
Than Tuberculosis
(MOTT)
Photochromogens (Runyon Group I)
Mycobacterium kansasii
Mycobacterium marinum
Scotochromogens (Runyon Group II)
Mycobacterium scrofulaceum
Mycobacterium xenopi
Mycobacterium szulgai
Mycobacterium gordonae

Mycobacteria Other
Than Tuberculosis
(MOTT)
Non-Photochromogens
Mycobacterium avium complex
Rapid Growers
Mycobacterium fortuitum group
Mycobacterium chelonae
Mycobacterium abscessus

MOTT Infections :
Most Common Causes1

Chronic bronchopulmonary disease (adults, cystic fibrosis

patients)
Mycobacterium kansasii
M. avium complex
M. abscessus
M. xenopi (less frequent)
M. szulgai (less frequent)

Cervical or other lymphadenitis (especially children)

Mycobacterium avium complex
M. scrofulaceum (less frequent)

Skin and soft tissue disease

Mycobacterium marinum
M. fortuitum
M. chelonae
M. abscessus

Brown-Elliott and Wallace, Jr., Mandell, Douglas, and Bennett 2005

MOTT Infections :
Most Common Causes1

Disseminated infection (HIV-positive)

Mycobacterium avium complex
M. kansasii
Disseminated infection (HIV-negative)
Mycobacterium abscessus
Mycobacterium chelonae
Catheter associated infections
Mycobacterium fortuitum
M. abscessus
M. chelonae

Brown-Elliott and Wallace, Jr., Mandell, Douglas, and Bennett 2005

Antitubercular Drugs

First line drugs

Rifampicin

Ethambutol

Inhibits biosynthesis of arabinogalactan (major

polysaccharide of cell wall)

Isoniazid

Broad spectrum, bactericidal

Mechanism of action :
binding bacterial RNA polymerase interferes
RNA synthesis

Inhibits mycolic acid biosynthesis (activated by

KatG gen)

Streptomycin

Aminoglycoside
Mechanism of action
Interferes protein synthesis, damage of cell
membrane, misreading/miscoding genetic
code

Multidrug resistant tuberculosis

(MDR TB)

Resistant for 2 main drugs

(Rifampicin & INH) w/ or w/o other
TB drugs
Incidence increase 2% per
annum (WHO Sept 2006
425.000 MDR new cases)
World MDR prevalence MDR
4.3%
Indonesia : ??
DRS (drug resistance surveilance) :

XDR-TB:
Extensive Drug
Resistant TB
XDR = MDR-TB plus resistance to
at least 3 of the 6 available
classes of second line drugs
Of 17,690 isolates in 16 countries
during 2000-2004 (by CDC & WHO),
20% were MDR and 2% were XDR
XDR found in:
USA: 4% of MDR
Latvia: 19% of MDR
S Korea: 15% of MDR
From : TB Laboratory training course for Asia-Pasific held by WHO, Beijing

Upaya Penanggulangan TB

Awal th 1990-an WHO dan IUALTD

telah mengembangkan strategi
penanggulangan TB yang dikenal
sebagai strategi DOTS (Directly
Observed Treatment Short-course)
dan telah terbukti sebagai strategi
yang secara ekonomis paling efektif
(cost-effective)

Strategi DOTS
1.
2.

3.

4.
5.

Komitmen Politis
Pemeriksaaan Dahak Mikroskopis yang
Terjamin Mutunya
Pengobatan Jangka Pendek yang standar
bagi semua kasus TB dengan tatalaksana
kasus yang tepat, termasuk pengawasan
langsung pengobatan
Jaminan ketersediaan OAT yang bermutu
Sistem pencatatan dan pelaporan yang
mampu memberikan penilaian terhadap
hasil pengobatan pasien dan kinerja
program secara keseluruhan

Pedoman Nasional
Penanggulangan TB (2008)
Tatalaksana Pasien Tuberkulosis
1.
Penemuan pasien TB
2.
Diagnosis TB paru dan TB ekstra paru
3.
Klasifikasi Penyakit dan Tipe Pasien
4.
Pengobatan TB
5.
Tatalaksana TB Anak
6.
Pengawasan Menelan Obat
7.
Pemantauan dan Hasil Pengobatan TB
8.
Pengobatan TB pada Keadaan Khusus

Jenis, Sifat dan Dosis OAT

Jenis OAT

Sifat

Isoniazid (H)

Bakterisid

Dosis harian Dosis mingguan

(mg/kg)
(mg/kg)
5 (4 6)

10 (8 12)

Rifampicin (R) Bakterisid

10 (8 12)

10 (8 12)

Pirazinamid
(Z)

Bakterisid

25 (20 30)

35 (30 40)

Streptomycin
(S)

Bakterisid

15 (12 18)

--

Ethambutol
(E)

Bakteriostatik

15 (15 20)

30 (20 35)

sekian
dan
terima kasih