Nitrate reductase assay using sodium nitrate for rapid detection of multidrug resistant tuberculosis

Abstract

We validated the nitrate reductase assay (NRA) for the detection of multidrug-resistant Mycobacterium tuberculosis (MDR-TB) using sodium nitrate (NaNO 3 ) in replacement of potassium nitrate (KNO 3 ) as nitrate source. NaNO 3 is cheaper than KNO 3 and has no restriction on use which facilitates the implementation of NRA to detect MDR-TB. tuberculosis to reduce nitrate to nitrite, which is easily detected in a colored reaction (5). NRA was initially standardized using potassium nitrate (KNO 3 ) (1) as nitrate source, but due to its higher cost and restriction of use in several countries, some authors have used sodium nitrate (NaNO 3 ) as nitrate source (3, 6, 7). However, there is no study comparing the accuracy of NRA using NaNO 3 in replacement of KNO 3 . In this study, we evaluated the use of the NaNO 3 in the NRA test to detect MDR-TB in comparison with KNO 3 and results were compared to those obtained with the conventional proportion method (PM) performed on Löwenstein-Jensen (LJ) medium.One-hundred and six M. tuberculosis strains belonging to the collection of the Institute of Tropical Medicine of Antwerp, Belgium, were studied. The strains were cultured in LJ medium and incubated at 37°C for three weeks. The PM was performed according to Canetti et al (2), using critical concentrations of 0.2 μg/mL for INH and 40 μg/mL for RIF. The NRA was performed according to the previously reported methodology

Fast detection of Mycobacterium tuberculosis in culture-positive sputum samples by nitrate reductase activity

“…The addition of sodium nitrate in the medium caused fast bacillus phenotypic identification without impairing its growth, corroborating studies by (Meyer et al, 2003;Goh, Rastogi, 2010). Macedo et al (2012) reported that, when sodium and potassium nitrate were compared, the two were metabolized as substrate.…”

“…The 7H9 medium was prepared according to manufacturer's instructions, with 0.2% glycerol and supplemented with 10% Oleic acid, Albumin, Dextrose and Catalase -OADC (Becton Dickinson Diagnostic Systems, Sparks, MD, USA), 0.1% casein and a lyophilized mixture of antimicrobials -MGIT PANTA (Becton Dickinson Diagnostic Systems, Sparks, MD, USA). The modified media, named 7H9-N and OK-N, were prepared as described above with the addition of 1 mg/mL of sodium nitrate (CAAL, São Paulo SP Brazil) in their formulation (Macedo et al, 2012).…”