DIAGNOSIS OF

TUBERCULOSIS
Emerging Trends
Dr.T.V.Rao, MD.

Dr.T.V.Rao

Robert Koch Discovers

Mycobacterium

Dr.T.V.Rao

A Global Emergency
The Tuberculosis in the
beginning of the 21st Century
declared as Global Emergency
(WHO)
Dr.T.V.Rao

Why Tuberculosis is a Important

Disease.
Tuberculosis continues to be a
Important communicable disease.
A leading cause of morbidity and
mortality in Developing world.
Most Important communicable
disease in Bangladesh, China,
Indonesia, Africa, and Pakistan.

But it is Curable Disease

Dr.T.V.Rao

Tuberculosis is a Global Problem

Dr.T.V.Rao

Tuberculosis - Important
communicable disease spread by
Respiratory route

Dr.T.V.Rao

Why Everybody Concerned.

Tuberculosis kills young adults.

Premature death of the infected a
prominent future.
Today many are co infected with HIV.
The open cases of Tuberculosis infects a
few around his/her environment.
A social burden to the family, society and
Nations.
Dr.T.V.Rao

Tuberculosis in the era of

HIV / AIDS.
HIV / AIDS epidemic led to large
increase of Smear negative pulmonary
tuberculosis which in turn has led to
poor treatment out comes, and early
mortality
Frequently involves Lower lobes of
Lungs.
Dr.T.V.Rao

Why we fail to Diagnose

Tuberculosis.

Lack of health infrastructure.

Control is plagued with lack of
Accurate,
Robust,
and Rapid
Diagnostic methods,
Technologies.
Dr.T.V.Rao

Why we failed ( Cont )

Diagnostic services are poor, and so we
failed at Individual and community
levels.
Patients are diagnosed late.
Many patients are never diagnosed
before death.
Early deaths are burden to Social
Infrastructure and Economic loss.

Dr.T.V.Rao

Importance of Clinical services

Early diagnosis rests with clinicians,
whose contribution is immense in
prompt treatment.
A clinicians knowledge, proper
documentation are immense help in
Developing countries.
Dr.T.V.Rao

When to suspect Tuberculosis

Cough longer than 3 weeks.
Fever for 1 month, or both.
Blood stained sputum.
Night sweats, weight loss
Age between 14 and 70 years

( Correlates National Tuberculosis

Programme ).

Dr.T.V.Rao

DIAGNOSTIC METHODS

Dr.T.V.Rao

Diagnosis.

Tuberculosis is a diversified disease.

Any organs can be involved.
Any age group, gender no bar for
Tuberculosis.
Involvement of Lungs contribute to
majority of tuberculosis.
And involvement of Lungs is designated as
Pulmonary tuberculosis.
Dr.T.V.Rao

Diagnosis of Pulmonary
Tuberculosis

Majority of Adults suffer with pulmonary

tuberculosis.
Microbiological examination of Sputum
continues to be a Gold standard in proving
the Diagnosis.
Sputum examination in Children is not
sensitive in Diagnosis.
Radiological examination of Lungs, most
commonly prescribed investigation.
Dr.T.V.Rao

X - ray examination of chest most

easily available Investigation.

Dr.T.V.Rao

Microbiological
Investigations are essential
for definitive Diagnosis of
Tuberculosis.

Dr.T.V.Rao

Importance of Optimal Specimens

Pulmonary Tuberculosis is the
commonest presentation of Tuberculosis
Sputum is the Most important specimen
for identification and isolation of Acid
fast bacilli.
The developing countries suffers the most
important step in getting an ideal
sample.
Dr.T.V.Rao

Sputum specimens
Train the staff to obtain the appropriate
specimen
A few minutes of education to patients on
importance of ideal sample make a great
difference and improves the Diagnosis.

Dr.T.V.Rao

Observe to identify Sputum from

Saliva.
SPUTUM
Specimens appear mucoid
even, blood stained.
Contains many
Polymorphoneutrophils.

Dr.T.V.Rao

SALIVA
Appears clear, watery,
and frothy.
Contains many
squamous epithelial
cells
Absence of
Polymorphoneutrophils.

Role of Microscopy in Tuberculosis.

Microscopy for Diagnosis of Tuberculosis is

initiated in 1880
The conceptions have not changed since
then.
Best efforts should be put to obtain
sputum,
Processing of saliva loses all valuable clues
to diagnose.
Dr.T.V.Rao

Microscopy and Tuberculosis

Microscopy with
Ziehl Neelsens
staining
A century old
procedure

Dr.T.V.Rao

Why Microscopy
Only we need Microscope, and few
stains.
Most rapid, economical,
Can detect bacterial load.
A Diagnostic, and Prognostic tool.
A little of sputum 0.2 l is adequate.
A prompt diagnosis after searching
as few as 100 fields.

Dr.T.V.Rao

Limitation of Microscopy for

Tuberculosis.

Repeated sample examinations. load on

technical staff.
Training and dedication of Microscopist.
The load of bacilli must be more than
10,000 / 1 ml of sputum.
Low in sensitivity < 50 %
Repeated requests for samples
High drop out by patients, for repeated
samples.
Not dependable in pediatric age group.
Dr.T.V.Rao

Smear showing Acid Fast Bacilli.

Dr.T.V.Rao

What is Smear Positivity

WHO

All patients who have submitted

two
Specimens and found to be
positive
for identification of AFB
Dr.T.V.Rao

Processing Direct smear negative

specimens

Sputum Microscopy can be improved with

Sputum liquefaction, concentration and
gravity sedimentation.
Popular solvents
Sodium hypochlorite.
Sodium hydroxide.
Ammonium sulphate
N-acetyl-L-cysteine sodium
hydroxide.
Dr.T.V.Rao

Benefits of Liquefaction and

Concentration

Major studies showed processing of sputum

with chemicals and centrifugation improved
sensitivity up to 18 %.
Incremental yield ( positive with bleach
minus positives with Ziehl Neelsen stain)
up to 9 %.
Treating specimens with Sodium
hypochlorite is Mycobactericidal and also
kills HIV and improves the safety and
acceptability by technical staff.
Dr.T.V.Rao

When Microscopy fails

Smear negative tuberculosis.
In HIV infected patients, on many
occasions prove negative. in spite of
presence of bacilli, ( as few bacilli are
expectorated).
Needs concentration and liquefaction
with chemicals.
Time consuming, needs more
technical manpower

Dr.T.V.Rao

Growing role of
Fluorescent Microscopy

There is a growing need for screening for

AFB by Florescent Microscopy.
Several studies prove, Florescent
Microscopy in Diagnosis of Tuberculosis is
a priority,
Developing world should opt and initiate
florescent microscopy.
Dr.T.V.Rao

Acid Fast Bacilli as seen under

Fluorescent Microscope

Dr.T.V.Rao

Why we need Florescent

Microscopy

Useful when few bacilli are present.

Increases the sensitivity in HIV patients with
tuberculosis.
Reduces the time needed for testing.
About 15 times as many fields of view can be
scanned by fluorescent microscopy than by Ziehl
Neelsenmethod in the same period.
Increases the sensitivity by 10 %
Better conclusions with one or two specimens,
unlike Ziehl Neelsens method needing 3 or > 3
specimens.
Dr.T.V.Rao

Culturing Mycobacterium
Culturing for isolation of
Mycobacterium spp continues to be a
Gold standard, particularly in
Developing countries.
Need only 10 100 bacilli / 1 ml of
sputum.

Dr.T.V.Rao

Culturing
Most useful in

Surveillance,
Drug sensitivity testing patterns.
Identify treatment failures.
Useful in Patients presenting with
respiratory symptoms, X- rays
suggestive, but smear negative. Can prove
culture positive.
Cultures remain suggestive and helpful in
early treatment periods, failed drug
regimes.
Dr.T.V.Rao

Methods of Culturing.

Culturing on Lowenstein Jensons

culture medium remain the
affordable ,economical method in
developing world.

Dr.T.V.Rao

Limitation in Culturing
Mycobacterium spp are slow
growing.
Need 6 8 weeks for growing.
Specimens can be contaminated
while growing, needs repeated
specimens, in turn patients loose
confidence in Laboratories.

Dr.T.V.Rao

Recent facts on Culturing

Useful in HIV infected patients with
Tuberculosis.
As even few bacilli can be grown in
spite of smear negativity.
But the specimens to be incubated
for longer time as few bacilli are
present.

Dr.T.V.Rao

Pitfalls in Culturing

Specificity is lost due to

contamination.
Can yield false positive results in 1
4 % of the cases.
Cultures may be negative in spite of
x rays are suggestive of tuberculosis.

Dr.T.V.Rao

Growth of Acid fast bacilli on

L J Medium.

Dr.T.V.Rao

ADVANCES IN
CULTURING
TECHNIQUES.
There are emerging Modern Media
with accurate detection, are
replacing the Egg and Agar based
medium.

Dr.T.V.Rao

Emerging methods in Culturing

MGIT Mycobacterium growth
incubator tube method.
Growth occurs in shorter than egg
medium.
Usefulness in HIV patients established.
Contamination is less
But expensive to people in Developing
world.

Dr.T.V.Rao

Blood culturing for Mycobacterium

Useful in HIV patients, and children.
Effective in isolation of Atypical
mycobacterium.
But not cost effective.
May be important tool in future for
diagnosing Tuberculosis in HIV
infected.

Dr.T.V.Rao

Molecular Methods in
Diagnosis of Tuberculosis
Several methods are
available, mainly used as
Research tools
Dr.T.V.Rao

Real Time PCR replacing older

Methods

Dr.T.V.Rao

PCR How useful to our Patients?

PCR ( Polymerase chain reaction ) used by

several investigators.
However most cases can be diagnosed
with simple methods if effectively used.
The definite role of PCR continues to be
controversial
Above all not cost effective to Developing
countries.
Dr.T.V.Rao

Rapid Diagnostic Methods

in Tuberculosis
Past decade has seen several
emerging technologies
How far practicable ?
Dr.T.V.Rao

Emerging Rapid Methods.

1. Fast Plaque TB uses phage amplification
technology.
2. ELISA ( QuantiFERON TB )
3. Enzyme-Linked immunospot
( ELISPOT )
ELISPOT proved highly useful to detect active
tuberculosis in Adults and children.
Dr.T.V.Rao

Emerging Technology
MODS

Microscopic observation drug susceptibility

assay. ( MODS )
A new method gained importance in
several reviews.
Use a tissue culture plate based assay with
use of Middle Brook 7HG.
Needs a inverted light microscope.
Even the drug resistance can be tested
with Rifampicin,and Isoniazid.
Safe to work with cultures.
Dr.T.V.Rao

Non Specific Tests

Tuberculin test
( Mantoux Test )

Dr.T.V.Rao

Tuberculin Test
( Mantoux Test )

Test to be interpreted
in relation to clinical
evaluation.
Even the induration of
5 mm to be
considered positive
when tested on HIV
patients.
Lacks specificity.

Dr.T.V.Rao

Serology in Tuberculosis.
Several serological methods were
evaluated.
But never gained the acceptance of
the majority of the clinicians.
Serological tests are low sensitivity.
Many physicians depend on serology
in extra pulmonary tuberculosis.

Dr.T.V.Rao

Dealing with Tuberculosis

in HIV / AIDS patients.
Diagnosing Tuberculosis in
HIV infected is a priority and
improve quality of Life

Dr.T.V.Rao

HIV/AIDS - Tuberculosis

Consider the HIV status

Identify the severity of Tuberculosis.
Early use of chest radiography.
Maximal number of sputum smear
examinations.
Sputum concentration methods to be
encouraged even by smaller laboratories.
Explore the use of Florescent Microscopy.
All smear negative specimens should be
cultured.
Dr.T.V.Rao

Limitations of Rapid Tests

The testing needs advanced and
sophisticated infrastructure.
These tests are known for their
inability to diagnose between active
disease and latent infection.
Exclusively used in Developed
nations.

Dr.T.V.Rao

Extra pulmonary
Tuberculosis
Poses several challenges, Yet
no optimal, specific diagnostic
methods

Dr.T.V.Rao

Extra pulmonary Tuberculosis

A real challenge to Clinicians and

Laboratories.
Optimal specimen collection a priority,
Molecular Methods are growing need.
Clinicians start drug regimes on empirical
basis.
Several serological tests for antibody
determinations are evaluated.
Dr.T.V.Rao

Identification of Atypical
Mycobacterium
A growing concern on infections with
less known, uncommon Mycobacterium
in immunosupreesed, an emerging
infectious disease problem

Dr.T.V.Rao

Atypical Mycobacterium
Needs the help of reference
laboratories.
Needs different drug regimes, unlike
typical Mycobacterium isolates.
Now a gowning concern in the era of
AIDS.

Dr.T.V.Rao

Future perceptions

It is highly essential to explore and discover

rapid, simple, and accurate tuberculosis
diagnostic tools.
A massive investment, greater scientific interest,
political commitment a top priority,
Man power development, Human resource
utilization a greater concern.
Microscopy and Florescent Microscopy utilization
should be immediate concern, and strengthening
of treatment initiation protocols.
Effective methods in diagnosing smear negative
patients a growing priority.
Dr.T.V.Rao

Microscopy in Tuberculosis
TODAY
In spite of several
scientific, and
molecular advances
Microscopy in
Tuberculosis
continues to be
back bone in
Diagnosis.
Dr.T.V.Rao

Specific detection of active

Tuberculosis cases in
patients with HIV infection or
AIDS is feasible and
improves the rate of early
diagnosis and successful
treatment of Tuberculosis.
Dr.T.V.Rao

Dr.T.V.Rao

Created for Medical graduates and

Health care workers in Developing world
Dr.T.V.Rao, MD.
e mail; doctortvrao@gmail.com
Dr.T.V.Rao