TRADITIONAL TESTS AND RECENT DIAGNOSTIC MODALITIES FOR TUBERCULOSIS WITH EMPHASIS TO MOLECULAR DETECTION TECHNIQUES, DRUG SENSITIVITY ASSESMENT IN INDIAN PERSPECTIVE
5. ADVANTAGES OF MGIT
• all type of specimens
• continuously monitored
• positive signals over 10-12 days
• non-radiometeric
• cheaper than BACTEC
6. DST using rapid culture method
MODS
• microscopic-observation drug-susceptibility assay
• antituberculous drug containing Middle brook
7H9 broth
• growth (cord formation) detected using inverted
light microscope
• time to detection- 7 days
• sensitivity 95% & specificity 100%
7. IFN-γ release assays
• in vitro assays
• whole blood incubated with TB antigens
• measures interferon (IFN-γ) released by
sensitized T cells
• measures immune reactivity to M.tb
9. QuantiFERON TB Gold
• combines IFN-γ release
technology with the
diagnostic power of
synthetic TB-specific
peptides (ESAT-6 and
CFP-10) to provide the
best available method
of diagnosing TB
infection
• FDA-approved for the
detection of LTBI, 2001.
10. Analysis of IGRA report
Positive IGRA
• person has been infected with TB bacteria.
additional tests are needed to determine if
the person has latent TB infection or TB
disease
Negative IGRA:
• latent TB infection or TB disease is not likely
11. MOLECULAR METHODS
• uncultivable or difficult to culture
• HIV infected patients
• rapid result
• quantitative information (viral load)
• susceptibility testing (drug resistance) without
culture
12. • may not differentiate active infection as DNA
from a dead organism during antibiotic
treatment can be detected and amplified by
PCR
• High cost
13. Nucleic acid amplification tests
• amplify M. tuberculosis-specific nucleic acid
sequences using a nucleic acid probe
• sensitivity at least 80% in most studies
• Require 10 bacilli/ml of given sample
• specificity 98% to 99%.
14. Molecular methods used in NAAT
For amplification
• Polymerase chain reaction
• Transcription mediated amplification
• Loop mediated isothermal amplification
• Ligase chain reaction
• Real time PCR
For detection:
• Electrophoresis
• Line probe assay
• Real time detection
15. Xpert MTB/RIF
• detects M. tuberculosis & rifampicin
resistance conferring mutations
• unprecedented sensitivity-smear negative,
culture positive specimens
• provides results directly from the sputum
within 100 minutes
• Endorsed by WHO since 2011
17. Xpert MTB/RIF from different samples
SAMPLE SENSITIVITY SPECIFICITY
Sputum 88% 99%
Lymph node aspirate 84.9% 92.5%
CSF 79.5% 98.6%
Pleural fluid 43.7% 98.1%
Gastric lavage/aspirate 83.8% 98.8%
Source: WHO policy update for Xpert MTB/RIF(2013)
18. Line probe assays
• PCR/hybridization
technique to identify
Mycobacterium
Tuberculosis
• identify drug-resistant
strains by detecting the
most common single
nucleotide
polymorphorisms
associated with
resistance
20. • In-house PCR tests are widely used in the
developing countries
• The line probe assays (LPA) are suitable only
for national or regional level laboratories
because of its complexity and bio safety
requirements
21. Molecular DST- candidates?
• Patients suspected or at high risk of having drug-
resistant TB
• very ill patients
• HIV infected patients
• who do not get better while taking standard first-
line therapy
22. Lateral flow urine lipoarabinomannan
(LF-LAM)assay
• From urine of active TB patients
• Endorsed by WHO in 2015
• Only for HIV patients- seriously ill/ low CD4
count