OBJECTIVES: The purpose of this study was to develop and assess a rapid method for pyrazinamide resistance detection in Mycobacterium tuberculosis using nicotinamide in a colorimetric resazurin assay. METHODS: We have tested M. tuberculosis isolates using nicotinamide in a 96-well format with the redox indicator resazurin (REMA) and compared results using the BACTEC 460-TB system with two concentrations of pyrazinamide (100 and 300 mg/L), as well as the Wayne method for detecting pyrazinamidase activity. Mutations in the pncA gene were detected by DNA sequencing of the pyrazinamide-resistant strains. RESULTS: Out of 95 clinical isolates of M. tuberculosis tested, 25 were determined to be resistant by the Wayne, BACTEC (300 mg/L), and the REMA nicotinamide methods. Using a nicotinamide MIC>250 mg/L as the cut-off for defining resistance, only one strain was falsely labelled as resistant. The REMA nicotinamide assay demonstrated a sensitivity of 100% and a specificity of 98%. The BACTEC (100 mg/L) falsely classified 8 strains as resistant. DNA sequencing detected mutations in 18/22 of the pncA genes from pyrazinamide-resistant strains. CONCLUSIONS: The REMA plate using nicotinamide to detect resistance to pyrazinamide is a simple and rapid method that could be useful in limited-resource countries.
OBJECTIVES: The purpose of this study was to develop and assess a rapid method for pyrazinamide resistance detection in Mycobacterium tuberculosis using nicotinamide in a colorimetric resazurin assay. METHODS: We have tested M. tuberculosis isolates using nicotinamide in a 96-well format with the redox indicator resazurin (REMA) and compared results using the BACTEC 460-TB system with two concentrations of pyrazinamide (100 and 300 mg/L), as well as the Wayne method for detecting pyrazinamidase activity. Mutations in the pncA gene were detected by DNA sequencing of the pyrazinamide-resistant strains. RESULTS: Out of 95 clinical isolates of M. tuberculosis tested, 25 were determined to be resistant by the Wayne, BACTEC (300 mg/L), and the REMAnicotinamide methods. Using a nicotinamide MIC>250 mg/L as the cut-off for defining resistance, only one strain was falsely labelled as resistant. The REMAnicotinamide assay demonstrated a sensitivity of 100% and a specificity of 98%. The BACTEC (100 mg/L) falsely classified 8 strains as resistant. DNA sequencing detected mutations in 18/22 of the pncA genes from pyrazinamide-resistant strains. CONCLUSIONS: The REMA plate using nicotinamide to detect resistance to pyrazinamide is a simple and rapid method that could be useful in limited-resource countries.
Authors: Niuris C Mirabal; Sergio L Yzquierdo; Dihadenys Lemus; Mariela Madruga; Yoslaine Milián; Miguel Echemendía; Howard Takiff; Anandi Martin; Patrick Van der Stuyf; Juan Carlos Palomino; Ernesto Montoro Journal: J Clin Microbiol Date: 2010-06-16 Impact factor: 5.948
Authors: Parissa Farnia; Mohammad Reza Masjedi; Foroozan Mohammadi; Payam Tabarsei; Poopak Farnia; Ali Reza Mohammadzadeh; Parvaneh Baghei; Mohammad Varahram; Sven Hoffner; Ali Akbar Velayati Journal: J Clin Microbiol Date: 2007-12-19 Impact factor: 5.948
Authors: Ekaterina V Kurbatova; Joseph S Cavanaugh; Tracy Dalton; Eleanor S Click; J Peter Cegielski Journal: Clin Infect Dis Date: 2013-07-09 Impact factor: 9.079
Authors: M J Nasiri; F Fardsanei; M Arshadi; B Deihim; Farima Khalili; M Dadashi; M Goudarzi; M Mirsaeidi Journal: New Microbes New Infect Date: 2021-05-05