Molecular and phenotypic characterisation of Mycobacterium tuberculosis resistant to anti-tuberculosis drugs

Dr Cetrángolo Hospital, Buenos Aires, Argentina.

To characterise drug-resistant (DR), multidrug-resistant (MDR-) and extensively drug-resistant (XDR-) Mycobacterium tuberculosis isolates, and identify their genetic profiles, drug resistance levels and resistance-conferring mutations.

Phenotypic drug susceptibility testing methods were used to determine drug resistance profiles. Minimal inhibitory concentrations (MICs) of isoniazid (INH), rifampicin (RMP) and levofloxacin (LVX) from 169 DR tuberculosis (TB) isolates, 78 of them monoresistant to INH, 13 to RMP, 7 to LVX, and 71 MDR-TB, were determined. Multiplex allele-specific polymerase chain reaction and DNA sequencing were used to detect mutations in katG, rpoB and gyrA/B genes. Genotyping was performed using spoligotyping and insertion sequence 6110 restriction fragment length polymorphism.

In total, 38.9% of the INH-resistant (INH(R)) isolates had an MIC ≥ 32 g/ml; 61.3% of RMP-resistant (RMP(R)) isolates had an MIC ≥ 64 g/ml and 55.6% of the LVX-resistant (LVX(R)) isolates had an MIC 4 ≥ 16 g/ml. The main mutations found in INH(R) isolates were katG315 (53.7%) and inhAP-15 (25.5%), whereas in RMP(R) isolates the main mutations were rpoB531 (61.9%), followed by rpoB526 (16.7%). LVX(R) isolates showed mutations in gyrA94/90. Haarlem, LAM and T were the main spoligotyping families found. katG315 was mainly associated with Haarlem and LAM, whereas inhAP-15 was associated with T.

Several isolates showed an association between high INH(R) levels and katG mutation; others from the Haarlem family were prone to becoming MDR-TB and continue to circulate in the community.