Avaliação da qPCR para determinação de genes de Mycobacterium tuberculosis associados à resistência à Isoniazida e à Rifampicina em amostras de escarros de pacientes multibacilares com Tuberculose pulmonar

Abstract

Early detection of resistance to drugs used to treat tuberculosis (TB) from Mycobacterium tuberculosis isolates, represent a challenge to Tuberculosis Control Program (TCP), because most antimicrobial susceptibility testing needs bacillus isolation in growth medium. Molecular methods have greater possibilities for rapid determination of resistance, they aim to detect mutations in M. tuberculosis genes associated with antimicrobial resistance and have promising results for Isoniazid and Rifampin, which are the major drugs of basic therapeutic regimen for TB treatment. This study evaluated the performance of PCR assay in Real Time (qPCR), directly by sputum samples and cultivation of strains isolated from patients with pulmonary tuberculosis multibacillar from Polyclinic Cardoso Fontes. 171 samples were analyzed by qPCR, targeting genes: katG, inhA and rpoB and the results were compared using the Nitrate Reductase Method (MNR). The resistance frequency for the evaluated samples was 9.36 % for Isoniazid , 5.85% for Rifampin and 4.09 % for both drugs. The katG and rpoB genes were associated with resistance to Isoniazid (p = 0.0001, 95% CI 3.61 to 21.01) and to Rifampin (p = 0.0001, 95% CI 6.34 to 51.05), respectively . The inhA gene was not associated with resistance to Isoniazid. There was acceptable agreement between phenotypic assay, MNR, and qPCR (sputum samples) for mutation detection for katG genes (kappa index: 0.4097) and rpoB (kappa index: 0.4985). The selected genes katG and rpoB were associated with resistance to Isoniazid and Rifampacina respectively, although the qPCR has had underperformed the MNR in this study. However, the molecular tests continue as promising targets for achieving faster results. In this context , optimizations in molecular testing should be performed to allow faster and more reliable diagnostics for resistance to anti -TB drugs and thus provide an appropriate treatment for the patient and reduce the transmission of TB.