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Multidrug resistant tuberculosis: risk factors and the application of innovative techniques for the rapid identification of resistances

Risultati finali

A rapid, culture-independent, and technically simple method has been developed for the prediction of isoniazid, streptomycin, ethambutol resistance in mycobacteria directly from clinical specimen (e.g. sputum). Background: The diagnostic detection of drug resistances in mycobacteria is impeded by the slow growth of many species, e.g. Mycobacterium tuberculosis. Resistance to the first-line therapeutic of these drugs can be predicted from specific mutations in resistance genes. Previous methods to detect these mutations were either too expensive and requiring special equipment, or little sensitive, requiring previous cultures. Key innovative features: Resistance-specific mutations are detected by PCR-RFLP, requiring only a PCR machine and a simple (minigel-) electrophoresis chamber. The technique is amenable to be marketed in a diagnostic kit format. Benefits: Any diagnostic laboratory with minimal technical equipment will be able to diagnose isoniazid resistance within 48 hours. This can be done without having to culture mycobacteria for which a special license and specific laboratory safety equipment would be needed. Current state of the art: The diagnostic detection of drug resistances in mycobacteria is impeded by the slow growth of many species, e.g. Mycobacterium tuberculosis. Resistance to the first-line therapeutic drug streptomycin can be predicted from specific mutations in resistance genes. Previous methods to detect these mutations were either too expensive and requiring special equipment, or little sensitive, requiring previous cultures. The improvement over the current state of the art is that resistance-specific mutations can be detected in a routine diagnostic setting by any microbiology laboratory, requiring only a PCR machine and a simple (minigel) electrophoresis chamber. The technique is amenable to be marketed in a diagnostic kit format.