Background: Rifampicin resistance in M. tuberculosis involves mutations in the rpoβ gene, encoding the β subunit of RNA polymerase. Objective: To identify the rpoβ gene mutations in strains of M. tuberculosis isolated from the Health Sub-Region in North Lima, Perϊ. Materials and methods: 73 sputum samples from patients with pulmonary tuberculosis were cultured on Lφwenstein-Jensen media. 62 samples, with more than 10 colonies per tube, were analyzed for susceptibility to isoniazid, streptomycin, rifampicin and ethambutol. DNA was isolated, amplified by PCR, followed by the cloning of the amplified product in the vector pGEM-T, transformation of competent E. coli bacteria, selection of recombinant clones and sequencing of the plasmid DNA for the determination of the polymorphisms in the rpoβ gene. Results: 52 strains out of 73 isolates (83,9%) were resistant to rifampicin (Rif r ), while 10 isolates (16,1%) were found to be rifampicin susceptible (Rif s ). 51 out of 52 Rif r strains contained mutations in the rpoβ gene associated with rifampicin resistance. 20 mutations were identified. The most frequently observed mutations were localized in the codons Ser-531 (62,7 %), His-526 (15,72%), Asp-516 (11,78%) and Gln-513 (5,88 %). No mutations were detected in the 10 Rif s strains. 94,2% of the Rif r strains were also resistant to isoniazid. Conclusions: rpoβ gene mutations associated with rifampicin resistance were present in almost all Rif r strains analyzed. Almost all Rif r strains were also resistant to isoniazid.
Antecedentes: La resistencia a rifampicina en M. tuberculosis involucra mutaciones en el gen rpoβ que codifica a la subunidad β de la ARN polimerasa. Objetivo: Identificar las mutaciones del gen rpoβ , en cepas de M. tuberculosis asociadas con resistencia a rifampicina aisladas de la Subregiσn de Salud Lima Norte, Perϊ. Materiales y métodos: Se cultivó en Lowestein - Jenseen 73 muestras de esputo de pacientes con tuberculosis pulmonar. A 62, con más de 10 colonias por tubo, se les comprobó susceptibilidad a isoniazida, rifampicina, estreptomicina y etambutol. Se realizó la extracción de ADN por PCR, clonación en el vector pGEM-T, transformación, selección de clonas recombinantes y secuenciamiento del ADN plasmídico para la determinación de los polimorfismos del gen rpoβ. Resultados: 52 (83,9%) cepas fueron resistentes a rifampicina (Rif r) y 10 (16,1%) susceptibles (Rif s ). Se encontró alteraciones en el gen rpoβ en 51 de 52 cepas Rif r Se identificaron 20 mutaciones. Las mutaciones mαs frecuentes fueron encontradas en los codones Ser-531 (62,7%), His-526 (15,7%), Asp-516 (11,8%) y Gln-513 (5,9%). No se observσ mutaciσn alguna en las 10 cepas Rif s. 94,2% de nuestras cepas Rif r fueron también resistentes a isoniazida. Conclusiones: Se encontraron mutaciones en el gen rpoB de casi todas las cepas Rif r ; asimismo, casi todas las cepas Rif r fueron también resistentes a isoniazida.