Sputum PCR-single-strand conformational polymorphism test for same-day detection of pyrazinamide resistance in tuberculosis patients

J Clin Microbiol. 2009 Sep;47(9):2937-43. doi: 10.1128/JCM.01594-08. Epub 2009 Jun 17.


Pyrazinamide is a first-line drug for treating tuberculosis, but pyrazinamide resistance testing is usually too slow to guide initial therapy, so some patients receive inappropriate therapy. We therefore aimed to optimize and evaluate a rapid molecular test for tuberculosis drug resistance to pyrazinamide. Tuberculosis PCR-single-strand conformational polymorphism (PCR-SSCP) was optimized to test for mutations causing pyrazinamide resistance directly from sputum samples and Mycobacterium tuberculosis isolates. The reliability of PCR-SSCP tests for sputum samples (n = 65) and Mycobacterium tuberculosis isolates (n = 185) from 147 patients was compared with four tests for pyrazinamide resistance: Bactec-460 automated culture, the Wayne biochemical test, DNA sequencing for pncA mutations, and traditional microbiological broth culture. PCR-SSCP provided interpretable results for 96% (46/48) of microscopy-positive sputum samples, 76% (13/17) of microscopy-negative sputum samples, and 100% of Mycobacterium tuberculosis isolates. There was 100% agreement between PCR-SSCP results from sputum samples and Mycobacterium tuberculosis isolates and 100% concordance between 50 blinded PCR-SSCP rereadings by three observers. PCR-SSCP agreement with the four other tests for pyrazinamide resistance varied from 89 to 97%. This was similar to how frequently the four other tests for pyrazinamide resistance agreed with each other: 90 to 94% for Bactec-460, 90 to 95% for Wayne, 92 to 95% for sequencing, and 91 to 95% for broth culture. PCR-SSCP took less than 24 hours and cost approximately $3 to $6, in contrast with the other assays, which took 3 to 14 weeks and cost $7 to $47. In conclusion, PCR-SSCP is a relatively reliable, rapid, and inexpensive test for pyrazinamide resistance that indicates which patients should receive pyrazinamide from the start of therapy, potentially preventing months of inappropriate treatment.

Publication types

  • Comparative Study
  • Evaluation Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Antitubercular Agents / pharmacology*
  • DNA, Bacterial / genetics
  • Drug Resistance, Bacterial*
  • Humans
  • Microbial Sensitivity Tests / economics
  • Microbial Sensitivity Tests / methods*
  • Mycobacterium tuberculosis / drug effects
  • Mycobacterium tuberculosis / genetics*
  • Polymerase Chain Reaction / methods
  • Polymorphism, Single-Stranded Conformational*
  • Pyrazinamide / pharmacology*
  • Sensitivity and Specificity
  • Sputum / microbiology
  • Time Factors
  • Tuberculosis / microbiology*


  • Antitubercular Agents
  • DNA, Bacterial
  • Pyrazinamide