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Journal of Biotechnology and Phytochemistry| Volume: 2
October 25-26, 2018 | Frankfurt, Germany
Joint Event
Biotechnology & Medical Microbiology
World Congress on
3
rd
International Conference on
Food Science & Technology
Early detection of multidrug resistant (MDR)
Mycobacterium tuberculosis
in a single tube with in-
house designed fluorescence resonance energy transfer (FRET) probes using real-time PCR
Devendra Singh Chauhan
National JALMA Institute for Leprosy & other Mycobacterial Diseases (ICMR), India
R
apid and correct diagnosis is crucial for the management
of multidrug resistance (MDR) in
Mycobacterium
tuberculosis
(MTB). The present study aims at rapid diagnosis
for identification of multidrug resistance tuberculosis (MDR-TB)
using real-time PCR. FRET hybridization probes targeting most
prominent four selected codons for rpoB526 and 531 and for
katG314 and 315 genes were designed and evaluated on 143
clinical MTB isolates and paired sputa for rapid detection of
MDR-TB. The results of real-time PCR were compared with gold
standard L-J proportion method and further validated by DNA
sequencing. Of the 143MTBpositive cultures, 85 and 58 isolates
were found to be ‘MDR’ and ‘pan susceptible’, respectively by
proportion L-J method. The sensitivity of real-time PCR for the
detection of rifampicin (RIF) and isoniazid (INH) were 85.88 and
94.11%, respectively, and the specificity ofmethodwas found to
be 98.27%. DNA sequencing of 31 MTB isolates having distinct
melting temperature (Tm) as compared to the standard drug
susceptible H37Rv strain showed 100% concordance with real-
time PCR results. DNA sequencing revealed the mutations at
Ser531Leu, His526Asp of rpoB gene and Ser315Thr, Thr314Pro
of katG gene in RIF and INH resistance cases. This real-time PCR
assay that targets limited number of loci in a selected range
ensures direct and rapid detection of MDR-TB in Indian settings.
However, future studies for revalidation as well as refinement
are required to break the limitations of MDR-TB detection.
Speaker Biography
Devendra Singh Chauhan is a scientist in the department of Microbiology and Molecular
biology at National JALMA Institute for Leprosy & other Mycobacterial Diseases(ICMR). His
interest and researches are towards investigation of
Mycobacterium Tuberculosis
and has
successfully worked on the various results of the same. He also worked as a core scientist
in group of Dr. V M Katoch, Former secretary DHR & DG, ICMR (Indian Council of Medical
Research).
e:
devchauhan01@yahoo.co.in