rpoB gene high-resolution melt curve analysis: a rapid approach for diagnosis and screening of drug resistance in tuberculous meningitis

Diagn Microbiol Infect Dis. 2015 Oct;83(2):144-9. doi: 10.1016/j.diagmicrobio.2015.06.010. Epub 2015 Jun 18.

Abstract

Timely and rapid diagnosis of multidrug resistance in tuberculous meningitis (TBM) is a challenge both for a microbiologist and neurologist. The present study was conducted to evaluate role of real-time polymerase chain reaction (PCR) using rpoB, IS6110, and MPB64 as targets in diagnosis of TBM in 110 patients and subsequent high-resolution melt (HRM) curve analysis of rpoB gene amplicons for screening of drug resistance. The sensitivity of smear, culture, and real-time PCR was 1.8%, 10.9%, and 83.63%, respectively. All 120 control patients showed negative results. With HRM rpoB analysis, rifampicin resistance was detected in 3 out of 110 cases of TBM (3.33%). Subsequently, results of HRM analysis were confirmed by rpoB gene sequencing, and mutations were observed at 516 (2 patients) and 531 (1 patient) codons, respectively. rpoB HRM analysis can be a promising tool for rapid diagnosis and screening of drug resistance in TBM patients in 90minutes.

Keywords: Diagnosis; High resolution melt curve; Resistance; Tuberculous meningitis.

Publication types

  • Comparative Study
  • Evaluation Study

MeSH terms

  • Adult
  • DNA-Directed RNA Polymerases / genetics*
  • Drug Resistance, Bacterial*
  • Humans
  • Mass Screening / methods
  • Mycobacterium tuberculosis / classification
  • Mycobacterium tuberculosis / drug effects*
  • Mycobacterium tuberculosis / genetics
  • Mycobacterium tuberculosis / isolation & purification*
  • Real-Time Polymerase Chain Reaction / methods*
  • Sensitivity and Specificity
  • Time Factors
  • Transition Temperature*
  • Tuberculosis, Meningeal / diagnosis*
  • Tuberculosis, Meningeal / microbiology

Substances

  • DNA-Directed RNA Polymerases
  • RNA polymerase beta subunit