This study shows a high level of agreement of these three low-cost methods compared with the PM for rapid detection of rifampicin and isoniazid resistance. However, more standardization is needed for ethambutol and streptomycin using the MTT test and resazurin microtitre assay. The nitrate reductase assay might represent an inexpensive procedure for rapid detection of resistance to first-line drugs in low-resource countries.

These three simple methods might become inexpensive alternative procedures for rapid detection of rifampicin resistance in low-resource countries.

No abstract

The nitrate reductase assay (NRA) was used as an alternative method for detection of resistance to the first-line antituberculous drugs isoniazid, rifampicin, ethambutol and streptomycin. A total of 320 strains of Mycobacterium tuberculosis were studied and the results compared with the proportion method (PM) on Lö wenstein-Jensen medium. The mean time to obtain results was 10 days and the overall agreement between the NRA and PM was 98?8 %. The NRA was easy to perform and represents a useful tool for rapid and accurate determination of drug-resistant M. tuberculosis strains in low-resource countries.

The direct detection of pyrazinamide resistance in Mycobacterium tuberculosis is sufficiently difficult that many laboratories do not attempt it. Most pyrazinamide resistance is caused by mutations that inactivate the pyrazinamidase enzyme needed to convert the prodrug pyrazinamide to its active form. We evaluated two newer and simpler methods to assess pyrazinamidase activity, the nitrate reductase and malachite green microtube assays, using nicotinamide in place of pyrazinamide. A total of 102 strains were tested by these methods and the results compared with those obtained by the classic Wayne assay. Mutations in the pncA gene were identified by sequencing the pncA genes from all isolates in which pyrazinamide resistance was detected by any of the three methods. Both the nitrate reductase and malachite green microtube assays showed sensitivities of 93.75% and specificities of 97.67%. Mutations in the pncA gene were found in 14 of 16 strains that were pyrazinamide resistant and in 1 of 4 strains that were sensitive by the Wayne assay. Both of these simple methods, used with nicotinamide, are promising and inexpensive alternatives for the rapid detection of pyrazinamide resistance in limited-resource countries.