Detection And Characterization Of Mutations In Rifampicin Resistant Mycobacterium Tuberculosis Clinical Isolates By Dna Sequencing

摘要

Background: Multiple Drug Resistant Tuberculosis (MDR-TB) is increasing because of widespread application and resultsin selection of mutants resistant to other components of short course chemotherapy. Resistance to Rifampicin can beconsidered as a surrogate marker for MDR-TB and the target gene for detection of rifampicin resistance is the rpo gene.Aims: To detect and characterize mutations in the rpo B region of Rifampicin resistant isolates of Mycobacteriumtuberculosis by automated DNA sequencing.Methods: Absolute concentration method was used to determine the MIC of Rifampicin for 44 M. tuberculosis isolates(21 respiratory, 3 ocular, 3 cerebrospinal fluid and 17 biopsies). Automated DNA sequencing was performed in the ABI310 Genetic Analyser.Results: Five isolates (2 sputa and one each from bronchoalveolar lavage, lymph node and endometrial biopsies) wererifampicin resistant with MIC greater than 128 mg/ml. Three of the five isolates showed mutations. Two of the isolateshad the common missense mutation at codon 531(Ser?Leu), the other isolate showed three insertions and two of themdid not show any mutation in the sequenced rpo B region.Conclusions: DNA sequencing technique is a rapid, conclusive and more advantageous technique than theconventional susceptibility testing for detection of rifampicin resistance in terms of the risk involved and timeconsumption.