Paula Fernanda Gonçalves dos Santos1, Elis Regina Dalla Costa2,7+, Daniela M Ramalho1, Maria Lucia Rossetti2,3, Regina Bones Barcellos1,2,7, Luciana de Souza Nunes4, Leonardo Souza Esteves2, Rodrigo Rodenbusch2, Richard M Anthony5, Indra Bergval5, Sarah Sengstake5, Miguel Viveiros6, Afrânio Kritski1,7, Martha M Oliveira1,7
1Universidade Federal do Rio de Janeiro, Faculdade de Medicina, Programa Acadêmico de Tuberculose, Programa de Pós-Graduação em Clínica Médica, Rio de Janeiro, RJ, Brasil
2Fundação Estadual de Produção e Pesquisa em Saúde, Centro de Desenvolvimento Científico e Tecnológico, Porto Alegre, RS, Brasil
3Universidade Luterana do Brasil, Porto Alegre, RS, Brasil
4Universidade Federal do Rio Grande do Sul, Centro de Biotecnologia, Programa de Pós-Graduação em Biologia Celular e Molecular, Porto Alegre, RS, Brasil
5Royal Tropical Institute, KIT Biomedical Research, Amsterdam, The Netherlands
6Universidade Nova de Lisboa, Instituto de Higiene e Medicina Tropical, Unidade de Microbiologia Médica, Global Health and Tropical Medicine, Lisboa, Portugal
7Rede Brasileira de Pesquisa em Tuberculose, Rio de Janeiro, RJ, Brasil
BACKGROUND To cope with the emergence of multidrug-resistant tuberculosis (MDR-TB), new molecular methods that can routinely be used to screen for a wide range of drug resistance related genetic markers in the Mycobacterium tuberculosis genome are urgently needed.
OBJECTIVE To evaluate the performance of multiplex ligaton-dependent probe amplification (MLPA) against Genotype®MTBDRplus to detect resistance to isoniazid (INHr) and rifampicin (RIFr).
METHOD 96 culture isolates characterised for identification, drug susceptibility testing (DST) and sequencing of rpoB, katG, and inhA genes were evaluated by the MLPA and Genotype®MTBDRplus assays.
RESULTS With sequencing as a reference standard, sensitivity (SE) to detect INHr was 92.8% and 85.7%, and specificity (SP) was 100% and 97.5%, for MLPA and Genotype®MTBDRplus, respectively. In relation to RIFr, SE was 87.5% and 100%, and SP was 100% and 98.8%, respectively. Kappa value was identical between Genotype®MTBDRplus and MLPA compared with the standard DST and sequencing for detection of INHr [0.83 (0.75-0.91)] and RIFr [0.93 (0.88-0.98)].
CONCLUSION Compared to Genotype®MTBDRplus, MLPA showed similar sensitivity to detect INH and RIF resistance. The results obtained by the MLPA and Genotype®MTBDRplus assays indicate that both molecular tests can be used for the rapid detection of drug-resistant TB with high accuracy. MLPA has the added value of providing information on the circulating M. tuberculosis lineages.
Financial support: CNPq/MCT (Process CNPq/INCT-TB 573548/2008-0, 478033/2009-5), FAPERJ (E: 26/110974/2011).
MV was supported by project “Ciência sem Fronteiras/Professor Visitante Especial” (CAPES/MEC/Brazil - Nº 88881.064961/2014-01).
Received 19 August 2016
Accepted 21 February 2017