RESUMO
INNO-LiPA Mycobacteria (LiPA; Innogenetics, Zwijnaarde, Belgium) is a kit for the simultaneous detection and identification of Mycobacterium species in culture and identifies the Mycobacterium tuberculosis complex, the M. avium complex (MAC), and the following Mycobacterium species: M. kansasii, M. avium, M. intracellulare, M. scrofulaceum, M. gordonae, M. xenopi, and the M. chelonae-M. abscessus complex. The assay, which targets the 16S-23S rRNA spacer region, was evaluated on 157 mycobacterial strains that had been identified by conventional techniques and PCR-restriction enzyme analysis of the hsp65 gene (PRA). Forty-seven reference strains consisting of 37 different species and 110 human clinical isolates were submitted to the test, and all were hybridized with the Mycobacterium genus probe (MYC) on the LiPA strip (100% sensitivity). Ninety-four isolates hybridized to their corresponding species- or complex-specific probes; only one isolate phenotypically identified as M. gordonae did not react with its specific probe (99.4% accuracy). Thirty-seven MAC strains were phenotypically identified to the complex level and to the species level by LiPA as M. avium (n = 18) or M. intracellulare (n = 7) or as belonging to the M. avium-M. intracellulare-M. scrofulaceum complex (n = 12). Of the last 12 strains, 10 had M. avium PRA patterns and 2 had M. intracellulare PRA patterns. Three isolates that had been identified as a single species by conventional identification were proven to be mixed cultures by the LiPA assay. The whole procedure can be performed in 1 working day, starting with the supernatant of a small amount of bacterial mass that had been treated by freezing and then boiling.
Assuntos
Proteínas de Bactérias , DNA Espaçador Ribossômico/genética , Infecções por Mycobacterium/microbiologia , Mycobacterium/classificação , RNA Ribossômico 16S/genética , RNA Ribossômico 23S/genética , Kit de Reagentes para Diagnóstico , Chaperonina 60 , Chaperoninas/genética , Sondas de DNA , Humanos , Mycobacterium/genética , Hibridização de Ácido Nucleico/métodos , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Padrões de Referência , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Especificidade da Espécie , Fatores de TempoRESUMO
In order to study polymorphisms of the DNA insertion sequence 6110 (IS6110) in Mycobacterium tuberculosis strains isolated from Colombian patients, together with resistance to antituberculous medications in the Department of Quindío, Colombia, a prospective study was conducted using a consecutive sample of 59 patients with symptomatic pulmonary tuberculosis whose cases had been confirmed by bacilloscopy, both with and without a history of treatment. The patients, who were participating in the Tuberculosis Control Program of the Regional Health Institute of Quindío in Armenia, included all individuals attending local health centers and hospitals between March and July 1993 who were referred to the regional institute. Sputum specimens from each patient were cultured and subjected to drug sensitivity tests. Subsequently, restriction fragment length polymorphisms (RFLP) of IS6110 from 27 patients were analyzed. The patients' treatment histories were used to classify their cases according to WHO criteria. Forty-five cultures were found positive, 44 for M. tuberculosis and 1 for M. africanum. Initial drug resistance was observed in 4 of 42 new cases, or 9.5% (95% CI: 0.6, 18), 2 showing resistance to isoniazid (INH) and 2 to isoniazid plus streptomycin (INH-SM). Acquired resistance was observed in 2 of the 3 chronic cases and relapses, the bacteria being resistant to isoniazid, rifampicin, and streptomycin (INH-RM-SM) in one case and to isoniazid, ethambutol, rifampicin, and streptomycin (INH-EMB-RM-SM) in the other. In those 27 strains subjected to RFLP analysis, the number of copies of IS6110 ranged from 6 to 17. Similarity coefficients revealed five distinct groups of strains. Overall, the RFLP analysis permitted most of the strains to be distinguished from one another, implying that the polymorphisms involved are sufficient to permit effective employment of this technique, which appears to have considerable potential for use in epidemiologic studies and in work designed to provide a basis for tuberculosis control program decision-making.
Assuntos
DNA Bacteriano/genética , Mycobacterium tuberculosis/genética , Polimorfismo de Fragmento de Restrição , Tuberculose Pulmonar/microbiologia , Mapeamento Cromossômico/métodos , Colômbia/epidemiologia , Humanos , Estudos Prospectivos , Tuberculose Resistente a Múltiplos Medicamentos/genética , Tuberculose Pulmonar/epidemiologiaRESUMO
The purpose of this study was to determine the polymorphism of insertion segment 6110 (IS6110) in strains of Mycobacterium tuberculosis isolated from Colombian patients as well as the current status of resistance to antituberculosis drugs in the department of Quindío, Colombia. To this end, a prospective study was performed with a consecutive sample of 59 patients who sought care at local health centers and hospitals in rural and urban areas of Quindío from March to July 1993. The patients in the sample had symptomatic pulmonary tuberculosis confirmed by bacteriologic inspection of sputum, with and without a history of treatment, and were participants in the Tuberculosis Control Program of the Sectional Health Institute of Quindío in Armenia, Colombia. Sputum cultures and drug sensitivity tests were done. Later, restriction fragment length polymorphisms (RFLP) of IS6110 were analyzed in accordance with the protocols of van Soolingen et al. (1992). Cases were classified by treatment history, applying the criteria of WHO (1991). The results showed 44 cultures positive for M. tuberculosis and one positive for M. africanum. Primary drug resistance was found in 4 of 42 cultures, or 9.5% (CI 95%: 0.6 to 18); 4.8% were resistant to isoniazid (INH) and 4.8% to isoniazid and streptomycin (INH-SM). Acquired resistance was found in two of three cultures, or 66% (to isoniazid, rifampicin, and streptomycin [INH-RM-SM] and to isoniazid, ethambutol, rifampicin, and streptomycin [INH-EMB-RM-SM]). In 27 strains submitted to RFLP analysis, the number of copies of IS6110 varied from 6 to 17. Similarity coefficients revealed five distinct groups.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Mycobacterium tuberculosis/genética , Polimorfismo de Fragmento de Restrição , Tuberculose Pulmonar/epidemiologia , Antituberculosos/uso terapêutico , Técnicas Bacteriológicas , Colômbia/epidemiologia , Resistência Microbiana a Medicamentos , Etambutol/farmacologia , Humanos , Isoniazida/farmacologia , Modelos Genéticos , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/crescimento & desenvolvimento , Estudos Prospectivos , Rifampina/farmacologia , População Rural , Estreptomicina/farmacologia , Tuberculose Pulmonar/tratamento farmacológico , População UrbanaRESUMO
Hemos examinado 148 muestras de suero prevenientes de enfermos sufriendo tuberculosis pulmonar(exámen directo del exputo positivo) y extrapulmonar (ganglionar). De esos mismos pacientes, la serología VIH fue efectuada. Según los resultados de la serología ELISA P32 y el antígeno sulfolipídico SLIV, la sensibilidad de la prueba donó valores muy bajos, de 48.6//para el antígeno P32 y 21.4//para el SLIV. La especificidad de la prueba fue de 76.9//para el antígeno P32