Detection of multidrug-resistant Mycobacterium tuberculosis directly from sputum samples of patients from Jakarta, Indonesia by radioisotope-based PCR-dot blot hybridization

Southeast Asian J Trop Med Public Health. 2012 Jan;43(1):89-95.

Abstract

The problem of eradicating tuberculosis (TB) has become more complicated by the emergence of multidrug resistant TB (MDR-TB). Any rapid laboratory method that can be used to detect drug susceptibility of Mycobacterium tuberculosis (MTB) is urgently needed. In this study, we employed the radioisotope (32P)-based PCR-dot blot hybridization method on sputum samples from patients in Jakarta, Indonesia. Bacterial DNA was extracted using BOOM method. KatG and rpobeta were amplified by PCR and katG315 or rpobeta531 mutations were identified by dot blot hybridization. Of 100 samples, 11% and 22% showed presence of mutation at codons 315 (AGC --> ACC) of katG and 531 (TCG --> TTG) of rpobeta, respectively. Five percent of the samples showed both mutations. This method is rapid, sensitive, and reliable and can be used to screen large numbers of samples in epidemiological studies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antitubercular Agents / pharmacology
  • Bacterial Proteins / analysis
  • Bacterial Proteins / genetics
  • Catalase / analysis
  • Catalase / genetics
  • Codon / genetics
  • DNA, Bacterial / analysis
  • DNA-Directed RNA Polymerases
  • Female
  • Humans
  • Immunoblotting / methods*
  • Indonesia / epidemiology
  • Male
  • Mutation / genetics
  • Nucleic Acid Hybridization / methods
  • Phosphorus Radioisotopes
  • Polymerase Chain Reaction / methods*
  • Sensitivity and Specificity
  • Sputum / microbiology*
  • Tuberculosis, Multidrug-Resistant / diagnosis*
  • Tuberculosis, Multidrug-Resistant / epidemiology
  • Tuberculosis, Multidrug-Resistant / genetics

Substances

  • Antitubercular Agents
  • Bacterial Proteins
  • Codon
  • DNA, Bacterial
  • Phosphorus Radioisotopes
  • rpoB protein, Mycobacterium tuberculosis
  • Catalase
  • katG protein, Mycobacterium tuberculosis
  • DNA-Directed RNA Polymerases