RESUMEN
SETTING: Cetrangolo Hospital, Vicente Lopez, Argentina, 1995-1999. OBJECTIVE: To describe a home-made reverse-line blot hybridisation assay for the detection of rifampicin resistance-associated mutations in the rpoB gene of Mycobacterium tuberculosis, and to evaluate the usefulness of this rifampicin oligonucleotide, or 'RIFO' assay, to predict rifampicin resistance. DESIGN: A total of 135 M. tuberculosis isolates from the Cetrangolo Hospital were tested using the RIFO assay, the proportion method and the Mycobacterial Growth Indicator Tube (MGIT 960). In addition, 120 drug-susceptible isolates from the Netherlands were included. RESULTS: The results obtained with the proportion method and the MGIT 960 system were in full agreement. In the RIFO assay, 90 of the 97 rifampicin-resistant isolates were correctly identified (sensitivity 92.8%, positive predictive value 100%). All of the drug-susceptible isolates were correctly predicted by the RIFO assay. CONCLUSIONS: With this home-made molecular test, rifampicin resistance in M. tuberculosis can be predicted in colonies isolated in culture in only 1 day, and can therefore shorten the laboratory turn around time for rifampicin susceptibility testing by weeks. In principle the test can also be applied directly to Zichl-Neelsen slides and clinical material, as has been demonstrated for another reverse-line blot-based assay for M. tuberculosis, spoligotyping.
Asunto(s)
Antibióticos Antituberculosos/uso terapéutico , ARN Polimerasas Dirigidas por ADN/genética , Mycobacterium tuberculosis/aislamiento & purificación , Hibridación de Ácido Nucleico/métodos , Rifampin/uso terapéutico , Tuberculosis Resistente a Múltiples Medicamentos/microbiología , Tuberculosis Pulmonar/microbiología , ADN Bacteriano/análisis , Humanos , Pruebas de Sensibilidad Microbiana/métodos , Mycobacterium tuberculosis/genética , Reacción en Cadena de la Polimerasa , Sensibilidad y Especificidad , Tuberculosis Resistente a Múltiples Medicamentos/tratamiento farmacológico , Tuberculosis Pulmonar/tratamiento farmacológicoRESUMEN
New technologies for rapid detection and drug susceptibility testing of mycobacteria are needed in clinical laboratories in order to attempt a rapid diagnosis and effective treatment of tuberculosis (TB) specially of cases resistant to isoniazid (H) and rifampin (RMP) (MDR-TB). A total of 218 pulmonary specimens from 132 HIV coinfected patients were processed and inoculated into the Mycobacterial Growth Indicator Tube system (MGIT, Becton Dickinson, MD) and on Lowenstein-Jensen (L-J) and Stonebrink (SB) solid media. The average time for recovering Mycobacterium tuberculosis (MTB) from MGIT was 18.3 days and 31.0 days from solid media. Of the patients 14.4% (19/132) were only diagnosed by MGIT. In another experiment susceptibility tests by the classical proportion method (PM) in L-J medium with H, RMP, streptomycin (SM), para-aminosalycilic-acid and ethambutol (EMB) were carried out on 120 isolates. The results were later compared with those obtained by determining the minimal inhibitory concentration (MIC) for each drug in MGIT against the above mentioned isolates. MIC results from MGIT method were available in an average of 5 days (3-10) and they correlated (correlation index 0.9974) with those of drug susceptibility obtained by the PM. A 4.7% (2/42) of disagreement among detecting isolates resistant to H and SM was found between PM and MGIT. Our results showed MGIT as a useful, safe and timesaving culture system, specially for detecting TB resistance. It might be used in clinical laboratories to improve the proper management of TB patients.