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Revista CON-CIENCIA

Print version ISSN 2310-0265

Abstract

MERCADO MICHEL, Brayan  and  VASQUEZ MICHEL, Aneth. Validation of the Real Time Polymerase Chain Reaction (qPCR) coupled to melting curves as an alternative tool for the diagnosis of Multi-Drug Resistant Tuberculosis. Rev.Cs.Farm. y Bioq [online]. 2021, vol.9, n.1, pp.77-96. ISSN 2310-0265.

ABSTRACT Introduction: Tuberculosis is an infectious-contagious disease, caused by various species of the Mycobacterium tuberculosis Complex, it is estimated that one third of the world population is affected by what represents a threat to public health, mainly by the emergence of multidrug-resistant strains (MDR-TB). In Bolivia, 7,538 people are reported sick with Tuberculosis, the latest data on MDR-TB indicate an increase of 0.2% per year, in 2018  there was 3.1% of MDR-TB. Currently in our country molecular methods are used to identify this infectious agent; however, there is very little or no work on the application of molecular methods for the precise and effective detection of MDR-TB strains that give validity to the results issued. This work resolves the question of whether real-time PCR (RT-qPCR) coupled to melting curves is an applicable alternative diagnostic tool for the identification of multidrug-resistant tuberculosis. Materials and methods: We worked with 74 strains of Mycobaterium tuberculosis phenotypically identified by culture (method of proportions, Canetti Rist) as a gold standar. The genetic material for molecular methods was obtained by the column assay, two primary controls were used for the determination of resistance to the drugs Isoniazid and Rifampicin, both the controls and the samples were processed by RT-qPCR coupled to melting curves, by means of temperature changes of dissociation.  Results: The diagnostic test parameters of the test demonstrated sensitivity: 67.4%, specificity: 83.3%, Accuracy: 73.97%, PPV: 85.3%, NPV: 64.1% for Isoniazid. While for Rifampicin: Sensitivity: 97%, Specificity: 20%, Accuracy: 58.9%, PPV: 55.4% and NPV: 87.5% Conclusion: The method evaluated for the determination of resistance to Isoniazid presents a balance between sensitivity and specificity, therefore it represents a reliable diagnostic alternative, while for resistance to Rifampicin it presents a high sensitivity that is very useful for endemic countries such as ours. .

Keywords : Tuberculosis; Multidrug-resistant tuberculosis; Real-time PCR coupled to melting curves; Validation.

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