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Tuberculosis (Edinb) ; 91 Suppl 1: S43-8, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22088321

RESUMO

Rapid tuberculosis (TB) detection is critical for disease control, and further quantitation of Mycobacterium tuberculosis (Mtb) in sputum is valuable for epidemiological and clinical studies. We evaluated a simple, robust and cost-efficient in-house DNA extraction and downstream Taqman approach for detection and quantitation of Mtb genomes from sputum of newly-diagnosed TB patients and non-TB controls. DNA was extracted using guanidine isothiocyanate and silica-based spin columns in less than 2 h, stored frozen, and Taqman assays were used to detect Mtb with IS6110 and quantify it targeting RD1 and IS1081. The Taqmans had a sensitivity >95% in 108 culture-confirmed TB patients and specificity of 100% in 43 non-TB controls. Genome counts were correlated with the Mycobacterial Growth Indicator Tubes' (MGIT) time-to-detection values (1/TTD × 1000; rho = 0.66; p < 0.001) in 91 TB patients (33 excluded with MGIT contamination). This linear relationship was nearly identical between mycobacteria isolated from sputum and H37Rv Mtb grown in-vitro to its log phase. TB treatment between 3 and 7 days was associated with lower 1/TTD × 1000 values but not with genome counts. Together, our protocol provides rapid, specific, inexpensive and quantitative detection of Mtb DNA in fresh or stored sputa making it a robust tool for prompt TB diagnosis, and with potential use for clinical and epidemiologic studies.


Assuntos
DNA Bacteriano/isolamento & purificação , Mycobacterium tuberculosis/isolamento & purificação , Escarro/microbiologia , Tuberculose Pulmonar/diagnóstico , Adulto , Estudos de Casos e Controles , Feminino , Genoma Bacteriano , Humanos , Masculino , Pessoa de Meia-Idade , Mycobacterium tuberculosis/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Sensibilidade e Especificidade , Manejo de Espécimes/métodos , Fatores de Tempo
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