Determination of the susceptibility of Mycobacterium tuberculosis to pyrazinamide in liquid and solid media assessed by a colorimetric nitrate reductase assay

Abstract

The CONRAS tests were rapid, cheap and easy to perform and interpret. Both tests should be evaluated on extended strain batteries in multicentre studies before they can be considered for use in susceptibility testing of M. tuberculosis to pyrazinamide in resource-limited settings.

Evaluation of Methods for Testing the Susceptibility of Clinical Mycobacterium tuberculosis Isolates to Pyrazinamide

“…Drug susceptibility testing in liquid media is costly, especially in some regions that do not have enough economic capabilities (6). Several other drug susceptibility-testing methods have been developed, including the molecular drug susceptibility test (mDST) based on the detection of a pncA mutation, the pyrazinamidase (PZase) activity assay, and colorimetric methods based on a minimal inhibitory concentration (MIC) or redox indicator (7,8,9,10,11,12). In this study, we used 432 clinical M. tuberculosis isolates to compare five methods for determination of the susceptibility of M. tuberculosis to PZA: the MGIT 960 system, the mDST, the PZase activity assay, and two colorimetric methods [the resazurin microtiter assay (REMA) and the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction test].…”

In vitro antimicrobial efficacy of a fixed-dose combination of RHZE against M. tuberculosis

“…This may have occurred because pyrazinamide has its great efficacy against microorganisms while at acidic pH, which was not observed in broth microdilution method. This occurred because broth microdilution method was used as a reference that proves that a cutoff value of 100 μg/mL of pyrazinamide revealed no changes in the results due to the pH (Syre, Ovrea, Grewal, 2010). Zhang, Zhang, Sun, (2003) proved that it is difficult for M. tuberculosis to maintain a neutral internal pH due to environmental conditions that cause an acidic pH.…”

“…Determination of the Minimum Inhibitory Concentration (MIC) of antimicrobials alone and in combination (R, H, Z, and E (Sigma ® )) was performed by the adapted BMM (NCCLS, 2003;Franzblau et al, 1998;Syre, Ovrea, Grewal, 2010). Rifampicin was dissolved in methanol and other drugs in sterile ultrapure water as specified by the manufacturer.…”

Antitubercular In Vitro Drug Discovery: Tools for Begin the Search

“…A choice more sensitive is the use of Alamar blue and resazurin assay, which changes from blue, nonfluorescent and oxidized form to pink and fluorescent upon reduction, can be read visual and fluorimetrically by exciting at 530 nm and detecting emission at 590 nm, and present high correlation with antitubercular gold standards methods (Collins & Franzblau, 1997). But, a more inexpensive colorimetric method, useful for evaluate antimycobacterial molecules in developing countries is using the ability that posses M. tuberculosis in to reduce nitrate to nitrite, nitrate reductase-based antibiotic susceptibility (CONRAS) test in liquid medium is perhaps the most cost-effective alternative for an ant i T B d r u g s c r e e n i n g program, with excellent results in comparison with other techniques, but is not useful for screening platforms that using nontuberculous mycobacteria nitrate negative (Kumar et al, 2005;Syre et al, 2010).…”