MUTATIONS IN THE RPOB GENE OF MYCOBACTERIUM TUBERCULOSIS IDENTIFIED BY SEQUENCING METHOD


Sureshbabu Ramalingam, Lakshmi Murali, Palaniswamy M

Abstract


ABSTRACT
Objective: To identify the mutation in the rpoB gene of Mycobacterium tuberculosis (MTB), using by sequencing method from pulmonary specimens
of presumptive TB patients belonging to the districts of Tamil Nadu.
Methods: A total of 8697 clinical specimens of presumptive MTB patients were collected from various districts of Tamil Nadu. Smear microscopy was
performed by light emitting diode fluorescent microscopy and all the smear positive samples were tested using line probe assay (LPA) to detect the
percentage of drug resistance pattern and to identify the missing mutation in LPA by the sequencing method.
Results: Among 4897 smear positives subjected to LPA method; 407 (8.3%) MTB was not detected and 16 (0.3%) showed invalid result; 4473 (91.4%)
strains showed MTB positive; 3695 (82.6%) were sensitive for both rifampicin (RIF) and isoniazid (INH) drugs; 502 (11.2%) were resistance for INH;
73 (1.6%) resistant for RIF; 203 (4.5%) were resistance for RIF and INH. Totally, 52 (1.2%) strains results cannot be confirmed by LPA and reported
as sensitive for RIF, because of the faint and the missing bands in both wild type and mutation. These strains were sequenced and 39 (75%) strains
showed resistant to RIF.
Conclusion: Hence LPA may be the molecular technology for the rapid, feasible and reliable method for the detection of multidrug resistant mutation
but few confusion bands cannot be reported as resistance, which should be confirmed by either conventional phenotypic drug susceptibility testing
or by sequencing method.
Keywords: Line probe assay, Sequencing, Mutation, Multidrug resistant tuberculosis.


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References


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Table 1: Total presumptive MDR cases tested, smear positive was diagnosed by LPA and the drug susceptibility results

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%, 39 strains reported as resistant)

MDR: Multidrug‑resistant, LPA: Line probe assay, MTB: Mycobacterium tuberculosis, RIF: Rifampicin, INH: Isoniazid

Table 2: 52 Strains were sequenced and the amino acid changes were observed in 39 strains, i.e., out of 52 strains 32 strains show the resistant in 531bp region and 7 strains were resistant in 526bp region in rpoB gene with 3 strains misread

Gene region

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Fig. 4: rpoB mutation-specific probes: MUT D516V, H526Y, H526D, S531L and rpoB wild type probes: WT1 to WT8

Asian J Pharm Clin Res, Vol 10, Issue 3, 2017, 382-387

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About this article

Title

MUTATIONS IN THE RPOB GENE OF MYCOBACTERIUM TUBERCULOSIS IDENTIFIED BY SEQUENCING METHOD

DOI

10.22159/ajpcr.2017.v10i3.16348

Date

01-03-2017

Additional Links

Manuscript Submission

Journal

Asian Journal of Pharmaceutical and Clinical Research
Vol 10 Issue 3 March 2017 Page: 382-387

Print ISSN

0974-2441

Online ISSN

2455-3891

Statistics

95 Views | 157 Downloads

Authors & Affiliations

Sureshbabu Ramalingam
KARPAGAM UNIVERSITY, COIMBATORE
India

Lakshmi Murali

Palaniswamy M


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