Rapid detection of multidrug-resistant Mycobacterium tuberculosis in Cotonou (Benin) using two low-cost colorimetric methods: resazurin and nitrate reductase assays Dissou Affolabi, 1,2 N’Dira Sanoussi, 1 Mathieu Odoun, 1 Anandi Martin, 2,3 Louis Koukpemedji, 1 Juan Carlos Palomino, 2 Luc Kestens, 4 Se ´ verin Anagonou 1 and Franc ¸ oise Portaels 2 Correspondence Dissou Affolabi affolabi_dissou@yahoo.fr 1 Laboratoire de Re ´ fe ´ rence des Mycobacte ´ ries, BP 817, Cotonou, Be ´ nin 2 Mycobacteriology Unit, Institute of Tropical Medicine, Antwerp 2000, Belgium 3 Me ´ decins Sans Frontie ` res, 75011 Paris, France 4 Immnunology Unit, Institute of Tropical Medicine, Antwerp 2000, Belgium Received 14 January 2008 Accepted 23 April 2008 We have evaluated two simple, rapid and low-cost colorimetric methods for the detection of multidrug-resistant Mycobacterium tuberculosis. A total of 151 M. tuberculosis strains were tested for resistance to rifampicin (RMP) and isoniazid by resazurin microplate assay (REMA) and nitrate reductase assay (NRA) in comparison with the conventional proportion method (PM) on Lo ¨ wenstein-Jensen medium. A complete agreement was found between NRA and PM, while one false RMP-susceptible result was found by REMA. REMA and NRA tests are rapid and inexpensive, and could be good alternatives to the conventional PM in low-resource countries. INTRODUCTION Tuberculosis (TB) is still a major public-health problem all over the world, particularly in developing countries. According to the latest World Health Organization (WHO) report in 2005, there were 8.8 million new TB cases and 1.6 million deaths were attributed to the disease worldwide (WHO, 2007). The situation becomes more complicated due to the rising human immunodeficiency virus/AIDS pandemic, the emergence of multidrug-resist- ant (MDR) TB and the recently described extensively drug- resistant TB (WHO, 2004; Aziz et al., 2006; Shah et al., 2007). Current standard methods for the detection of MDR Mycobacterium tuberculosis include the proportion method (PM) performed on Lo ¨wenstein–Jensen (LJ) medium or agar, absolute concentration and resistance ratio methods (Canetti et al., 1963, 1969; Kent & Kubica, 1985) and the radiometric method in the BACTEC-460 system (Roberts et al., 1983). However, these methods either are lengthy or produce radioactive waste that is difficult to manage in low-resource countries. Commercial methods, such as the mycobacteria growth indicator tube (MGIT) and molecular methods, have been introduced (Rossau et al., 1997; Palomino et al., 1999); however, though rapid, these methods are expensive and could not be easily implemented in developing countries. Recently, several rapid and inexpensive tests to detect drug resistance in M. tuberculosis have been developed (Wilson et al., 1997; Abate et al., 2004; Caviedes et al., 2000; Angeby et al., 2002; Palomino et al., 2002); unfortunately, such tests have been evaluated only in a few low-resource countries (Nateche et al., 2006; Rivoire et al., 2007) but not so far in a TB reference laboratory in West Africa. In this study we have evaluated two rapid and low-cost colorimetric tests, the nitrate reductase assay (NRA) and the resazurin microplate assay (REMA), for the detection of resistance to rifampicin (RMP) and isoniazid (INH) in strains of M. tuberculosis in a TB reference laboratory in Cotonou, Benin. The results were compared to those obtained by the standard PM performed on LJ medium. METHODS Setting. The study was performed in the National Reference Mycobacteriology Laboratory (Laboratoire de Re ´fe ´rence des Mycobacte ´ries) in Cotonou, Benin, a West African country. External quality control of the laboratory is performed by the Supranational Mycobacteriology Laboratory of the Institute of Tropical Medicine (ITM) in Antwerp, Belgium, and regular proficiency testing for drug susceptibility testing using the PM on LJ medium has shown excellent results. Abbreviations: INH, isoniazid; NRA, nitrate reductase assay; PM, proportion method; REMA, resazurin microplate assay; RMP, rifampicin; TB, tuberculosis; WHO, World Health Organization. Journal of Medical Microbiology (2008), 57, 1024–1027 DOI 10.1099/jmm.0.2008/000406-0 1024 2008/000406 G 2008 SGM Printed in Great Britain