A sputum PCR-SSCP test for same-day detection of pyrazinamide resistance in tuberculosis patients


Sheen, P; Mendez, M; Gilman, RH; Pena, L; Caviedes, L; Zimic, MJ; Zhang, Y; Moore, DA; Evans, CA; (2009) A sputum PCR-SSCP test for same-day detection of pyrazinamide resistance in tuberculosis patients. Journal of clinical microbiology. ISSN 0095-1137 DOI: 10.1128/JCM.01594-08

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Abstract

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 polymerase chain reaction 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 for sputum (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 sputa and 100% of Mycobacterium tuberculosis isolates. There was 100% agreement between PCR-SSCP results from sputa and Mycobacterium tuberculosis isolates and 100% concordance between 50 blinded PCR-SSCP re-readings by three observers. PCR-SSCP agreement with the four other tests for pyrazinamide resistance varied from 89-97%. This was similar to how frequently the four other tests for pyrazinamide resistance agreed with each other: 90-94% for Bactec-460; 90-95% for Wayne; 92-95% for sequencing; and 91-95% for broth culture. PCR-SSCP took less than 24-hours and cost approximately $3-$6 compared with the other assays that took 3-14 weeks and cost $7-$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.

Item Type: Article
Faculty and Department: Faculty of Infectious and Tropical Diseases > Dept of Immunology and Infection
Faculty of Infectious and Tropical Diseases > Dept of Clinical Research
Research Centre: Antimicrobial Resistance Centre (AMR)
TB Centre
PubMed ID: 19535526
Web of Science ID: 269439600035
URI: http://researchonline.lshtm.ac.uk/id/eprint/5190

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