Diagnosis of tuberculosis

Diagnosis ofDiagnosis of
tuberculosistuberculosis

Diagnosis of tuberculosisDiagnosis of tuberculosis
Active disease Latent tuberculosisActive disease Latent tuberculosis
1. Clinical suspicion,
2. Response to treatment,
3. Chest radiographs,
4. Staining for acid-fast bacilli
(AFB),
5. Culture for mycobacteria,
6. Nucleic acid amplification
(NAA) assays
1. Tuberculin skin test
2. Interferon γ

Active tuberculosisActive tuberculosis
An ideal test for active tuberculosis :An ideal test for active tuberculosis :
1. Rapid results (available within 1 day),
2. High sensitivity and specificity,
3. Low cost,
4. Robustness (ability to provide reproducible results
in a variety of settings),
5. Highly automated or easily performed without the
need for excessive sample preparation or technical
expertise,
6. Provide drug-susceptibility data.

6. The ideal test would be able to distinguish
between LTBI and active disease:
--For LTBI, such a test would distinguish true infection
from bacille Calmette-Guerin (BCG) vaccination
and infection with nontuberculous mycobacteria
(NTM).
--For active disease, it would be valuable to be able to
determine infectiousness, follow response to
therapy, distinguish Mycobacterium tuberculosis
from NTM in AFB positive specimens and obtain
drug-susceptibility information.

No ideal test for active TBNo ideal test for active TB
diagnosisdiagnosis

Clinical algorithm for the laboratoryClinical algorithm for the laboratory
diagnosis of tuberculosisdiagnosis of tuberculosis..

 Infection with TB generally requires
prolonged and close contact with a smear-
positive individual. Infection is influenced by
a variety of factors, including the number of
bacilli present in the sputum of the smear-
positive individual, the closeness of the
relationship, the duration of exposure, the age
of the contact (the risk is higher in children),
and the immunological status of the contact.

TB cannot be distinguished from other infections onTB cannot be distinguished from other infections on
the basis of clinical signs and symptoms alone.the basis of clinical signs and symptoms alone.
Most cases are characterized by an insidious, not very
alarming, and quite variable onset that depends on the
virulence of the causal pathogen, the patient’s age,
the organ infected, and the host’s immune status.
Symptoms can be classified into 2 groups:
1. Systemic symptoms. The most common systemic
symptoms are fever, loss of appetite, weight loss,
asthenia, profuse nocturnal sweating, and general
malaise.
2. Organ specific symptoms.

Pulmonary tuberculosisPulmonary tuberculosis
 To establish a diagnosis of pulmonary
tuberculosis, respiratory samples are submitted
to the laboratory for microscopy (AFB smear)
and for mycobacterial culture.

 Expectorated sputum is generally the starting
point. Three samples are collected on three
separate days and stained for AFB.
 Samples are generally sent simultaneously for
smear and culture, because culture data are
essential for confirmation of the diagnosis.

AcidAcid--fast stainingfast staining::
 The material to be examined on a glass slide andThe material to be examined on a glass slide and
staining with carbolstaining with carbol--fuchsin by means of either thefuchsin by means of either the
ZiehlZiehl--Neelsen or Kinyoun techniqueNeelsen or Kinyoun technique..
 The sensitivity of detection of organisms by acidThe sensitivity of detection of organisms by acid--fastfast
smears is increased if the specimen is digested andsmears is increased if the specimen is digested and
centrifuged and the concentrated sediment is stainedcentrifuged and the concentrated sediment is stained..
 Stained smears are usually examined under standardStained smears are usually examined under standard
brightbright--light microscopy with oil immersion, alight microscopy with oil immersion, a
technique that can be performed under verytechnique that can be performed under very
primitive conditionsprimitive conditions..

 The sensitivity of microscopic examination isThe sensitivity of microscopic examination is
low; the level of detection is approximatelylow; the level of detection is approximately
5000-10,000 bacilli per milliliter of secretions, if5000-10,000 bacilli per milliliter of secretions, if
100 oil immersion microscopic fields are100 oil immersion microscopic fields are
examinedexamined..
 In practice, 40% to 70% of patients withIn practice, 40% to 70% of patients with M.M.
tuberculosistuberculosis isolated in culture have positiveisolated in culture have positive
smears.smears.

The sensitivity tends to be highest in patients who
have cavitary disease and lowest in patients who
have weak cough or less advanced disease.

Fluorochrome staining procedureFluorochrome staining procedure ::
 Sensitivity of detection of acidSensitivity of detection of acid--fast organisms isfast organisms is
increased by a fluorochrome staining procedureincreased by a fluorochrome staining procedure
withwith auramine Oauramine O, a fluorescent stain, a fluorescent stain..
 This procedure requires use of a fluorescentThis procedure requires use of a fluorescent
microscope but is faster than acidmicroscope but is faster than acid--fast stainingfast staining
because the intensity of the fluorescent signalbecause the intensity of the fluorescent signal
enables slides to be scanned at lowerenables slides to be scanned at lower
magnificationmagnification..

In most situations in which an acidIn most situations in which an acid--fastfast
organism is detected by microscopy, itorganism is detected by microscopy, it
should be assumed to beshould be assumed to be MM.. tuberculosistuberculosis
until proven otherwise.until proven otherwise.

 If a patient is suspected of having pulmonary
tuberculosis but is smear negative on
expectorated sputum or is unable to produce
sputum for testing (30% of patients in one
series), further diagnostic testing may be
warranted.
 The options include:
1. Sputum induction (SI),
2. Fiberoptic bronchoscopy (FOB),
3. Gastric washings (GW) “limited role in adults”.

 The utility of FOB (or SI):
1. Generating a sample in patients who do not
have spontaneous sputum creates the potential
for making a diagnosis.
2. Rapid diagnosis (by positive smear or
histopathology) in either subset of patients
provides the potential for earlier intervention
and treatment while awaiting culture results.

Sputum induction
 Sputum induction provided appropriate samples
for diagnosis and increased the early diagnostic
yield significantly. Sputum induction also seems
to be cost-effective in the resource-poor setting.

Sputum induction
 Each successive sputum induction , up to three
in total, increased the yield for culture-positive
samples significantly. Sputum induction was
performed without difficulty in 96% of patients
and had an overall yield of 96.3% after three
tests, confirming the utility of repeated sputum
induction .

By contrast, the yield of fibrooptic
bronchoscopy was only 51.9%, making
sputum induction significantly more
sensitive.

Fibrooptic bronchoscopyFibrooptic bronchoscopy
 FOB encompasses BAL, bronchial brushings,
transbronchial biopsy, and postbronchoscopy sputum
collection.
The most productive use of FOB is in pulmonaryThe most productive use of FOB is in pulmonary
tuberculosis suspects who produce no sputum ortuberculosis suspects who produce no sputum or
who are smear negative and in patients in whomwho are smear negative and in patients in whom
there is considerable diagnostic uncertainty,there is considerable diagnostic uncertainty,
where lung biopsy may produce an alternativewhere lung biopsy may produce an alternative
diagnosis.diagnosis.

Fibrooptic bronchoscopyFibrooptic bronchoscopy
 Overall, cultures of M. tuberculosis from FOB
were positive in 90%. More significantly, a rapid
diagnosis was made in fully 72% of cases.
 The bronchial brushing smears was a result of
brushing caseous material in the bronchi when
visible and had a high diagnostic yield.

CulturesCultures
 Cultures of mycobacteria require only 10 to 100
organisms to detect M. tuberculosis.
 the sensitivity of culture is excellent, ranging
from 80% to 93%.
 the specificity is quite high, at 98%.
 Cultures increase the sensitivity for diagnosis of
M. tuberculosis and allow speciation, drug-
susceptibility testing, and, if needed, genotyping
for epidemiologic purposes

 There are three types of culture media:
1. Solid media, which includes
A. egg-based media (Lowenstein-Jensen)
B. agar-based media (Middlebrook 7H10 and
7H11),
2. Liquid media (Middlebrook 7H12 and other
broths).

Solid media:
 the standard.the standard.
 slower than liquid media.slower than liquid media.
 widely used alongside solid media to increasewidely used alongside solid media to increase
sensitivity and decrease recovery time.sensitivity and decrease recovery time.
 Lowenstein-Jensen 7H10 and 7H11 media mayLowenstein-Jensen 7H10 and 7H11 media may
detect mycobacteria in less than 4 weeks butdetect mycobacteria in less than 4 weeks but
they require incubation for as long as 6 to 8they require incubation for as long as 6 to 8
weeks before they can be classified as negative.weeks before they can be classified as negative.

Broth media:Broth media:
 broth media combined with DNA probes forbroth media combined with DNA probes for
rapid species identification typically providerapid species identification typically provide
results in less than 2 weeks with smear positiveresults in less than 2 weeks with smear positive
samples and somewhat longer with smearsamples and somewhat longer with smear
negative samples.negative samples.
 They allow more rapid determination of drugThey allow more rapid determination of drug
susceptibilities.susceptibilities.

Newer culture technologies:Newer culture technologies:
 TK MediumTK Medium uses multiple-color dye indicatorsuses multiple-color dye indicators
to identify M. tuberculosis rapidly.to identify M. tuberculosis rapidly.
 It can also be used for drug-susceptibility testingIt can also be used for drug-susceptibility testing
and can differentiate a contaminated specimen.and can differentiate a contaminated specimen.

Radiometric and Colorimetric DetectionRadiometric and Colorimetric Detection
SystemsSystems
 Radiometric culture systems (Radiometric culture systems (the BACTEC)the BACTEC)
incorporateincorporate 1414
C-labeled palmitic acid into a liquidC-labeled palmitic acid into a liquid
culture medium.culture medium.
 Growth of mycobacteria results in liberation ofGrowth of mycobacteria results in liberation of
1414
CO2 that can be measured by the detection device.CO2 that can be measured by the detection device.
The increased sensitivity of the system enablesThe increased sensitivity of the system enables
growth to be detected sooner, usually in 10 to 14growth to be detected sooner, usually in 10 to 14
days.days.
 A positive niacin production test or DNA probeA positive niacin production test or DNA probe
testing confirms an isolate as beingtesting confirms an isolate as being M. tuberculosisM. tuberculosis..

ChromatographyChromatography
 High-performance liquid chromatographyHigh-performance liquid chromatography
(HPLC) is a rapid and highly specific method for(HPLC) is a rapid and highly specific method for
detecting the unique pattern of mycolic acids fordetecting the unique pattern of mycolic acids for
identifying mycobacterial species.identifying mycobacterial species.
 Tuberculostearic acidTuberculostearic acid, a component of M., a component of M.
tuberculosis, can be easily detected even intuberculosis, can be easily detected even in
infinitesimal (femtomole) quantities by gas–infinitesimal (femtomole) quantities by gas–
liquid chromatography (GLC).liquid chromatography (GLC).

Nucleic Acid Amplification TestsNucleic Acid Amplification Tests
 In each test, either DNA or ribonucleic acidIn each test, either DNA or ribonucleic acid
(RNA) is amplified to detectable levels.(RNA) is amplified to detectable levels.
 Nucleic acid amplification tests include thoseNucleic acid amplification tests include those
that involve PCR amplification, transcriptionthat involve PCR amplification, transcription--
mediated amplification, strandmediated amplification, strand--displacementdisplacement
amplification, ligase chain reaction, and Q Betaamplification, ligase chain reaction, and Q Beta
replicase amplification.replicase amplification.
 NAA assays are also quite specific for M.
tuberculosis, with specificities in the range of
98% to 99%.

Two FDA-approved NAA assays are widely available for
commercial use:
1. The AMPLICOR M. tuberculosis uses DNA
polymerase chain reaction (PCR) to amplify nucleic
acid targets.
2. The Amplified Mycobaterium Tuberculosis Direct
(MTD) Test is an isothermal strategy for detection of
M. tuberculosis rRNA.
Both used on smear positive sample with a sensitivity of
70-90% and specificity of 93-99% (up to 100% in
some studies).

 The E-MTD (enhanced MTD test) brings with it an
improved sensitivity , especially in smear-
negative specimens.
 In smear negative patients, the sensitivity was
90.2%, and the specificity was 99.1%

 Amplification systems can detect as few asAmplification systems can detect as few as 1–101–10
organismsorganisms in clinical specimens and clearlyin clinical specimens and clearly
distinguishes M. Tuberculosis in the specimensdistinguishes M. Tuberculosis in the specimens
containing nucleic acids from human cells, othercontaining nucleic acids from human cells, other
mycobacterial species and common contaminatingmycobacterial species and common contaminating
organisms.organisms.
 These amplification techniques provide results in lessThese amplification techniques provide results in less
than a day and have been used for screening a variety ofthan a day and have been used for screening a variety of
clinical specimens.clinical specimens.
 The major limitation of NAA tests, they can not detectThe major limitation of NAA tests, they can not detect
drug sensitivity.drug sensitivity.

Polymerase chain reactionPolymerase chain reaction ((PCRPCR((
 PCR targets DNA, rRNA, insertion andPCR targets DNA, rRNA, insertion and
repetitive elements, and various protein-repetitive elements, and various protein-
encoding genes.encoding genes.
 The PCR amplification process can beThe PCR amplification process can be
completed in 2–4 h after obtaining thecompleted in 2–4 h after obtaining the
processed clinical sample. The detection assayprocessed clinical sample. The detection assay
requires an additional 2–24 h.requires an additional 2–24 h.

Disadvantages:Disadvantages:
1.1. PCR tests are currently expensive.PCR tests are currently expensive.
2.2. the PCR tests are unable to differentiatethe PCR tests are unable to differentiate
between viable and the dead AFB, which maybetween viable and the dead AFB, which may
lead to disagreement between the sputumlead to disagreement between the sputum
smear and PCR results.smear and PCR results.
3.3. PCR studies show unacceptable level ofPCR studies show unacceptable level of
sensitivity in smear negative but culture-sensitivity in smear negative but culture-
positive cases.positive cases.
False positive PCR results, usually due toFalse positive PCR results, usually due to
laboratory introduced contamination.laboratory introduced contamination.

 Effectiveness of the PCR technique inEffectiveness of the PCR technique in
diagnosing tuberculosis is related to:diagnosing tuberculosis is related to:
(a) DNA concentration in the clinical sample(a) DNA concentration in the clinical sample
(b) size of the target DNA sequence,(b) size of the target DNA sequence,
(c) repetitiveness of the amplified sequence,(c) repetitiveness of the amplified sequence,
(d) choice of primers, and(d) choice of primers, and
(e) expertise of the personnel conducting the assay.(e) expertise of the personnel conducting the assay.

TranscriptionTranscription--mediatedmediated
amplificationamplification ((TMATMA)) “MTD test”“MTD test”
 Isothermal, target-based amplification system isIsothermal, target-based amplification system is
based on amplification of ribosomal RNAbased on amplification of ribosomal RNA
(rRNA) unlike PCR, which is based on(rRNA) unlike PCR, which is based on
amplification of DNA.amplification of DNA.
 Probes that target rRNA are 10- to 100 foldProbes that target rRNA are 10- to 100 fold
more sensitive than those that target DNA.more sensitive than those that target DNA.

TranscriptionTranscription--mediatedmediated
amplificationamplification ((TMATMA((
 the amplified M. tuberculosis direct (MTD) testthe amplified M. tuberculosis direct (MTD) test
is entirely done entirely in one test tube, whichis entirely done entirely in one test tube, which
minimizes laboratory-introduced contamination.minimizes laboratory-introduced contamination.
 The entire test may be completed in 3–4 h afterThe entire test may be completed in 3–4 h after
obtaining the processed clinical specimen.obtaining the processed clinical specimen.
 But like PCR, the MTD test lacks sensitivity inBut like PCR, the MTD test lacks sensitivity in
the smear-negative but culture positive cases.the smear-negative but culture positive cases.

A reasonable use of NAA assays for rapidA reasonable use of NAA assays for rapid
diagnosis of pulmonary tuberculosis is as followsdiagnosis of pulmonary tuberculosis is as follows::
NAA assays should be used to confirm that a positive
AFB smear does indeed represent M. tuberculosis.
1. If both smear and NAA are positive, pulmonary
tuberculosis is diagnosed with near certainty.
2. If the smear is positive and the NAA is negative,
testing the sputum for inhibitors and repeating the
assay is warranted. If inhibitors are not detected, and
the process is repeated on additional specimens and is
negative, the patient can be presumed to have NTM

 If a sputum sample is smear negative (with
clinical suspicion is intermediate or high)) but E-
MTD “ enhanced Mycobaterium Tuberculosis
Direct” positive (only the E-MTD is approved
for smearnegative specimens), it is
recommended testing additional samples. If
further samples are E-MTD positive, the
patient can be assumed to have pulmonary
tuberculosis.

4. If both the smear and E-MTD are negative, an
additional specimen should be tested by E-
MTD. If negative, the patient can be assumed
not to have infectious pulmonary tuberculosis.

 NAA should not be performed on sputum
samples from cases in which the AFB smear is
negative and the clinical index of suspicion is
low.
 Testing should also be limited to those who
have not been treated recently for active disease.
((NAA tests are able to detect nucleic acids from bothNAA tests are able to detect nucleic acids from both
living and dead organisms and may be falsely positive).living and dead organisms and may be falsely positive).

Other NAA testsOther NAA tests
Strand displacement amplificationStrand displacement amplification ((SDASDA((
 SemiSemi--quantitative isothermal in vitro nucleic acidquantitative isothermal in vitro nucleic acid
amplification technique.amplification technique.
 The entire assay may be completed within four hoursThe entire assay may be completed within four hours
after the receipt of the processed clinical specimen.after the receipt of the processed clinical specimen.
 The amplification is carried out in a single tubeThe amplification is carried out in a single tube
(multiplexing), and the products can be differentiated(multiplexing), and the products can be differentiated
by the detection system, without significant loss ofby the detection system, without significant loss of
sensitivity.sensitivity.
 Species-specific SDA assays (for M. tuberculosis, M.Species-specific SDA assays (for M. tuberculosis, M.
avium, and M. kansasii) and a genus-specific assay haveavium, and M. kansasii) and a genus-specific assay have
also been developed.also been developed.

QQ--beta replicase amplificationbeta replicase amplification
 Q-Beta replicase enzyme is used for producingQ-Beta replicase enzyme is used for producing
RNA in the amplification reaction at a fixedRNA in the amplification reaction at a fixed
temperature.temperature.
 M. tuberculosis is detected with a sensitivity ofM. tuberculosis is detected with a sensitivity of
up to one colony-forming unit.up to one colony-forming unit.

Ligase chain reactionLigase chain reaction ((LCRLCR((
 LCR, a variant of PCR, is potentially useful forLCR, a variant of PCR, is potentially useful for
screening persons at high risk for developingscreening persons at high risk for developing
tuberculosis and extrapulmonary tuberculosis.tuberculosis and extrapulmonary tuberculosis.
 The LCx M. tuberculosis assay (Abbot), primarilyThe LCx M. tuberculosis assay (Abbot), primarily
makes use of the respiratory specimens. It has a highmakes use of the respiratory specimens. It has a high
sensitivity and specificity for smear-positive andsensitivity and specificity for smear-positive and
-negative specimens.-negative specimens.
 Currently, the use of this technique is limited by its highCurrently, the use of this technique is limited by its high
cost and lack of skilled personnel.cost and lack of skilled personnel.

Rapid detection of drug resistance
1.1. Line probe assays.Line probe assays.
2.2. Molecular beacons.Molecular beacons.
3.3. Phage amplification.Phage amplification.
4.4. Luciferase reporter phages.Luciferase reporter phages.

Pulmonary disease is suspectedPulmonary disease is suspected
1. Three induced or spontaneous (respiratory isolation) a.m.1. Three induced or spontaneous (respiratory isolation) a.m.
sputum samples for AFB staining and microscopy;sputum samples for AFB staining and microscopy;
- 5 to 10 mL each, accept watery specimens- 5 to 10 mL each, accept watery specimens
2. One sputum sample for MTb complex PCR;2. One sputum sample for MTb complex PCR;
3. Same three sputum samples for culture;3. Same three sputum samples for culture;
4. If effusion is present, consider sending diagnostic4. If effusion is present, consider sending diagnostic
thoracentesis sample for smear, culture, PCR, andthoracentesis sample for smear, culture, PCR, and
adenosine deaminase (ADA) testing- pleural biopsy hasadenosine deaminase (ADA) testing- pleural biopsy has
higher culture sensitivity;higher culture sensitivity;
5.Consider early morning gastric aspiration of 50 mL in5.Consider early morning gastric aspiration of 50 mL in
children or failed sputum inductions; and,children or failed sputum inductions; and,
6.Consider an IFN based assay on serum or Tuberculin Skin6.Consider an IFN based assay on serum or Tuberculin Skin
Test- negative test does not exclude disease.Test- negative test does not exclude disease.

Other than pulmonary diseaseOther than pulmonary disease
suspectedsuspected
1.Samples as you would for suspected pulmonary1.Samples as you would for suspected pulmonary
disease; and,disease; and,
2.Sample from suspected site. For tissue collections,2.Sample from suspected site. For tissue collections,
collect both with and without formalin.collect both with and without formalin.
a.a. Lymph nodeLymph node
-excisional biopsy preferred-excisional biopsy preferred
-fine needle aspiration if unable to obtain excisional-fine needle aspiration if unable to obtain excisional
biopsy, e.g. CT guided biopsy in setting of mesentericbiopsy, e.g. CT guided biopsy in setting of mesenteric
adenitisadenitis
-AFB smear and culture-AFB smear and culture
-histopathologic analysis-histopathologic analysis

suspected (contsuspected (cont.).)
b.b. CSFCSF
-cell count and differential-cell count and differential
- chemistries to include glucose and protein- chemistries to include glucose and protein
- AFB smear (typically negative) and culture (10 mL often- AFB smear (typically negative) and culture (10 mL often
required)required)
- consider PCR- consider PCR
c.c. Pleural, pericardial, or ascitic fluidPleural, pericardial, or ascitic fluid
-cell count and differential-cell count and differential
- chemistries to include glucose and protein- chemistries to include glucose and protein
-AFB smear and culture (lining biopsies have higher yield)-AFB smear and culture (lining biopsies have higher yield)
-consider ADA analysis-consider ADA analysis
-PCR-PCR

suspected (contsuspected (cont.).)
d.d. Other tissues (bone marrow, skin, liver, spleen, ..., kidney)Other tissues (bone marrow, skin, liver, spleen, ..., kidney)
- AFB smear and culture- AFB smear and culture
- histopathologic analysis, may consider specialized AFB- histopathologic analysis, may consider specialized AFB
staining, and subsequent targeted PCRstaining, and subsequent targeted PCR
-PCR-PCR
ee.. BloodBlood
- Culture - PCR- Culture - PCR
ff.. StoolStool, consider intestinal biopsy, consider intestinal biopsy
g.g. Urine (first morning mid-void stream collection), three dailyUrine (first morning mid-void stream collection), three daily
samplessamples
- urinalysis and conventional culture- urinalysis and conventional culture

Latent tuberculosisLatent tuberculosis

11..Tuberculin skin testTuberculin skin test
 Latent TB until very recently, has been diagnosed
exclusively by the tuberculin skin test (TST).
 The tuberculin skin test has both poor sensitivity and
specificity.
 The standard test: “the intracutaneous injection
(Mantoux's method) of 0.1 mL (5 TU) of PPD”
Tuberculin purified protein derivative (PPD)-RT23 with
Tween 80 administered at a dose of 2 tuberculin units
(TU) per 0.1 mL, which is biologically equivalent to the
recommended dose (5 TU) of the international
standard tuberculin (PPD-S).

Interpretation of Tuberculin Skin TestInterpretation of Tuberculin Skin Test
Reactions:Reactions:
≥≥5 mm:5 mm:
 Known or suspected HIV infection or otherKnown or suspected HIV infection or other
immunosuppressed condition immunosuppressed condition
 Recent close contact with infectiousRecent close contact with infectious
tuberculosis tuberculosis
 Chest film abnormalities suggestive of oldChest film abnormalities suggestive of old
tuberculosistuberculosis

Interpretation of Tuberculin Skin TestInterpretation of Tuberculin Skin Test
Reactions:Reactions:
≥≥10 mm10 mm
 Persons born in high-prevalence areasPersons born in high-prevalence areas
 Injection drug users Injection drug users
 Medically underserved, low-income populations: high-Medically underserved, low-income populations: high-
risk minority populations risk minority populations
 Residents of long-term care facilities (nursing homes,Residents of long-term care facilities (nursing homes,
correctional facilities, etc.) correctional facilities, etc.)
 Persons with medical conditions that increase the riskPersons with medical conditions that increase the risk
of tuberculosisof tuberculosis
≥≥15 mm15 mm All othersAll others

Potential Causes of FalsePotential Causes of False--Negative TuberculinNegative Tuberculin
Skin TestSkin Test
Factors Releated to the Person Being TestedFactors Releated to the Person Being Tested
 Concurrent infections Concurrent infections
 Human immunodeficiency virus Human immunodeficiency virus
 Other viral infections (measles, mumps, chickenpox) Other viral infections (measles, mumps, chickenpox)
 Bacterial diseases (typhoid fever, brucellosis, typhus, leprosy,Bacterial diseases (typhoid fever, brucellosis, typhus, leprosy,
pertussis, overwheiming tuberculosis) pertussis, overwheiming tuberculosis)
 Live virus vaccination (measles, mumps, polio)Live virus vaccination (measles, mumps, polio)
 Diseases affecting lymphoid organs (Hodgkin's disease,Diseases affecting lymphoid organs (Hodgkin's disease,
lymphoma, chronic lymphocytic leukemia, sarcoidosis)lymphoma, chronic lymphocytic leukemia, sarcoidosis)
 Immunosuppressive drugs (corticosteroids and many others)Immunosuppressive drugs (corticosteroids and many others)
 Age (newborns, elderly)Age (newborns, elderly)
 Metabolic conditions (advanced renal failure, malnutrition)Metabolic conditions (advanced renal failure, malnutrition)
 Recent surgeryRecent surgery
 BurnsBurns

Factors Related to the TestFactors Related to the Test
 Loss of antigen potency (exposure to heatLoss of antigen potency (exposure to heat
and/or light)and/or light)
 Incorrect administration (too little antigen, deepIncorrect administration (too little antigen, deep
injection)injection)
 Incorrect reading (inexperienced reader, readerIncorrect reading (inexperienced reader, reader
bias, recording error)bias, recording error)

2. Interferon2. Interferon γγ
 They include :They include :
1.1. QuantiFERONQuantiFERON--TB,TB,
2.2. QuantiFERONQuantiFERON--TB Gold,TB Gold,
3.3. T SPOTT SPOT--TB test.TB test.
 The basis of these tests is the detection in serum ofThe basis of these tests is the detection in serum of
either the release of IFN-either the release of IFN-γγ on stimulation ofon stimulation of
sensitized T cells by M. tuberculosis antigens insensitized T cells by M. tuberculosis antigens in
vitro (QuantiFERON) or detection of the T cellsvitro (QuantiFERON) or detection of the T cells
themselves (T SPOT-TB).themselves (T SPOT-TB).

QuantiFERONQuantiFERON--TB Gold:TB Gold:
 The ELISA-based QuantiFERON TB test and
the RD1 antigens, ESAT-6 (the early secreted
antigenic target 6) and CFP-10 (culture filtrate
protein-10), led to a more specific test.

T SPOTT SPOT--TB test: “TB test: “ELISPOT technique”
 In healthy BCG-vaccinated controls, the
ELISPOT was not confounded by BCG. In
TST-positive household contacts of a case of
active disease (expected cases of LTBI), 85%
were ELISPOT positive, suggesting a sensitivity
for LTBI of 85% if the TST is taken to be the
reference standard.

Diagnosis of tuberculosis

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