Genetic and phenotypic characterization of drug-resistant Mycobacterium tuberculosis isolates

The reemergence of Mycobacterium tuberculosis with increasing numbers of multi-drug resistant (MDR) strains has increased the need for rapid diagnostic methods.
Molecular bases of drug resistance have been identified for all of the main antituberculous drugs, and drug resistance results from changes in several target genes, some of which are still undefined. Drug resistance in M. tuberculosis is due to the acquisition of mutations in chromosomally encoded genes and the generation of multidrug resistance is a consequence of serial accumulation of mutations primarily due to inadequate therapy. Several studies have shown that resistance to isoniazid (INTI) is due to mutaions in kat G gene. The rpo B gene, which encodes the subunit of RNA polymerase, harbors a mutation in an 81 bp region in about 95% of rifampicin (RIF) reistant M. tuberculosis strains recovered globally. Streptomycin (STR) resistance is due to mutations in rrs and rpsl genes which encodes 16S SrRNA and ribosomal protein S12 respectively. Approximately 65% of clinical isolates resistant to ethambutol have a mutation in the embB gene.
Several susceptibility phenotypes methods have been developed. Among them were radiometric systems such as the BACTEC460 TB system reduces the test time considerably, but they still labor intensive, expensive and require manipulation of radioactive substances.
Rapid promising approach to determine drug resistant strains especially for rifampicin (RIF) is the use of mycobacteriophage. A number of low-cost colorimetric AST assays, such as the 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, the Alamar blue assay, and an assay based on microscopic detection of cord-like growth by M. tuberculosis, have been described. However, these tests have limitations; mycobacteria other than M. tuberculosis can produce cord factor, and INH can interfere with the formazan production in the MTT assay and give rise to false resistant results. Moreover, the use of a liquid medium in a microtiter plate format in these tests may be disadvantageous not only as a biohazard but also due to possible Contamination between wells.
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Mycobacterium Tuberculosis, Current status in Rapid Laboratory Diagnosis
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