INT J TUBERC LUNG DIS 9(8):901–906 © 2005 The Union Multicenter study of MTT and resazurin assays for testing susceptibility to first-line anti-tuberculosis drugs A. Martin,* N. Morcillo, † D. Lemus, ‡ E. Montoro, ‡ M. A. da Silva Telles, § N. Simboli, ¶ M. Pontino, † T. Porras, # C. León, # M. Velasco,** L. Chacon, †† L. Barrera, ¶ V. Ritacco, ¶ F. Portaels,* J. C. Palomino* * Institute of Tropical Medicine, Antwerp, Belgium; † Hospital Dr. A. Centrángolo, Vicente Lopez, Buenos Aires, Argentina; ‡ Instituto de Medicina Tropical ‘Pedro Kouri’, La Habana, Cuba; § Instituto Adolfo Lutz, Sao Paulo, Brazil; ¶ Institute Carlos G. Malbrán, Buenos Aires, Argentina; # Grupo de Micobacterias Subdirección de Investigación Instituto Nacional de Salud, Bogotá, Colombia; ** Instituto de Salud Publica de Chile, Santiago, Chile; †† Centro Nacional de SUMMARY Diagnostico y Referencia, Managua, Nicaragua OBJECTIVE: A multicentre evaluation was performed to assess two rapid low-cost methods, MTT (3-[4.5- dimethylthiazol-2-yl]-2.5-diphenyltetrazolium bromide) and resazurin assays, for testing the susceptibility of Myco- bacterium tuberculosis to the first-line anti-tuberculosis drugs rifampicin (RMP), isoniazid (INH), ethambutol (EMB) and streptomycin (SM). METHODS: Thirty coded M. tuberculosis strains were sent to seven laboratories located in Latin America, rep- resenting six countries. Each site performed the colori- metric assays, MTT and resazurin, blind for the first-line drugs RMP, INH, EMB and SM. The minimum inhibi- tory concentration results obtained were compared to the conventional proportion method on Löwenstein- Jensen medium. RESULTS: After establishing the breakpoint concentra- tions, excellent results were obtained for RMP, INH and EMB, with levels of specificity and sensitivity of between 96% and 99%. CONCLUSION: MTT and resazurin assays are promis- ing, accessible new alternative methods for middle- and low-resource countries that need low-cost methods to perform rapid susceptibility testing of M. tuberculosis to key anti-tuberculosis drugs. KEY WORDS: M. tuberculosis; MTT; resazurin; first-line drugs THE SPREAD of Mycobacterium tuberculosis strains resistant to the two key anti-tuberculosis (TB) drugs, isoniazid (INH) and rifampicin (RMP), defined as multidrug-resistant TB (MDR-TB), challenges the con- trol of the disease. 1,2 The emergence of MDR-TB high- lights the need for drug susceptibility testing (DST), both for proper patient management as well as for drug resistance surveillance, a priority defined by the World Health Organization (WHO) to detect areas of emerging resistance in time and to avoid dissemination of the disease. 3 In low-resource countries, where TB is endemic and MDR-TB is a more serious public health problem, DST is still currently based on conventional culture methods with solid media such as Löwenstein-Jensen (LJ) and Middlebrook agar, which are laborious and lengthy, requiring at least 4 weeks of incubation be- fore an isolate can be reported as susceptible or resis- tant. 4–6 Newer, more rapid culture methods in liquid media are increasingly being used, as they reduce the time needed for culture and DST. Due to its reliability and speed, the BACTEC TB-460 radiometric system (Becton Dickinson, Sparks, MD) was the first to achieve worldwide recognition. However, this method re- quires radioisotopes and an expensive machine 7,8 that are limiting factors for its implementation in low- resource countries. The commercial Mycobacterial Growth Indicator Tube (MGIT) system (Becton Dick- inson) is a good candidate to replace it; 9,10 it is rapid and reliable, but is still expensive in some low-resource countries. New molecular tools for rapid detection of drug resistance usually need skills, supplies, equipment and facilities that are not always available in endemic countries; in addition, as the molecular mechanisms of drug resistance are not completely understood, molecular methods fail to detect all those resistant strains in which no associated mutations have been identified to date. 11,12 Among the recently described new phenotypic methods for DST, rapid colorimetric methods, based on the ability of live bacteria to reduce an indicator and produce a change of visual colour, appear prom- ising techniques and have been described with success previously. 13–24 They are simple and require no so- Correspondence to: Anandi Martin, Mycobacteriology Unit, Institute of Tropical Medicine, Nationalestraat, 155, Antwerp, B-2000 Belgium. Tel: (132) 3 2476334. Fax: (132) 3 2476333. e-mail: amartin@itg.be Article submitted 5 November 2004. Final version accepted 27 December 2004.