Rapid detection of rifampicin-resistant Mycobacterium tuberculosis by in-house, reverse line blot assay.
Drug resistance in tuberculosis is a significant problem in countries endemic for tuberculosis. A sensitive, specific, and high-throughput reverse line blot assay (RLBA) for the detection of genotypic resistance to rifampicin (RIF) was designed and evaluated. DNA sequencing done for 72 resistant isolates from Delhi, for baseline data, showed mutations within the rpoB core region in all RIF-resistant strains. The RLBA includes oligonucleotide probes specific for wild-type and mutant sequences, allowing sensitive detection of both genotypes in a single assay. The assay based on reverse hybridization principle simultaneously detects 13 different mutations affecting 6 independent codons, including the most prevalent mutations at positions 531 and 526. Application of the method to a panel of 292 MDR TB isolates and susceptible strains from 5 different cities in India showed 98% concordance with the sequencing results. This rapid, simple, economical, and highly sensitive assay provides a practical alternative to sequencing for genotypic evaluation of RIF resistance in Mycobacterium tuberculosis.
Suresh
Naga
N
Department of Microbiology, All India Institute of Medical Sciences, New Delhi 110 029, India.
Singh
Urvashi Balbir
UB
Arora
Jyoti
J
Pande
Jitendra Nath
JN
Seth
Pradeep
P
Samantaray
Jyotish Chandra
JC
D000995:Antitubercular Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases; D024881:Drug Resistance, Bacterial; D006801:Humans; D015151:Immunoblotting / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics; D012293:Rifampin / Q000494:pharmacology*; D012680:Sensitivity and Specificity
D000995:Antitubercular Agents; D001426:Bacterial Proteins; C505805:rpoB protein, Mycobacterium tuberculosis; D012321:DNA-Directed RNA Polymerases; D012293:Rifampin