Rapid and accurate diagnosis of tuberculosis (TB) is the essential first step in the successful treatment and control of the disease in the individual and the community. In addition, rapid identification of drug- resistant TB is critical for tailoring effective regimens, particularly in areas with emerging drug-resistant TB. Current diagnostic tests for TB are cumbersome, expensive, slow and may lack sensitivity. Novel tests that provide an accurate result and that are affordable and feasible for studies of innovative diagnostic methods for detecting Mycobacterium tuberculosis in clinical specimens from clinical settings in a developing country, and to evaluate novel assays for the rapid diagnosis of drug- resistant TB. The purpose of this project is to develop and evaluate novel, rapid methods for the identification of M. tuberculosis and drug resistance from clinical specimens in a developing country setting. We will utilize the patient population of the University Hospital in Rio de Janeiro to perform pilot studies of candidate tests in patients suspected of having TB. Candidate tests will be evaluated in well-characterized patients for sensitivity, specificity, predictive value, and time required to provide results in comparison with conventional techniques. Candidate tests that show promise in the hospital setting will then be field tested in patients suspected of having TB at community clinics at the sites of our other trials (Project 1 and 2 of this proposal). Candidate tests to be developed and evaluated include: 1) C18-Carboxypropylbetaine (CB-18) specimen processing for enhancement of identification of M. tuberculosis by smear and in standard solid and radiometric media assays, as well as in nucleic acid amplification tests; 2) multiantigen serologic assays using lipid antigens of M. tuberculosis to distinguish active diseases from latent or absent infection; 3) solid phase amplification assays (e.g., Lineprobe (LIPA]) for rapid identification of both M. tuberculosis and specific resistance mutations; 4) use of immunocapture of BrdU-labeled mycobacterial DNA with PCR amplification to diagnose TB and identify phenotypic resistance after in vitro exposure to anti-mycobacterial drugs; and 5) use of redox dyes such as Alamar blue and triphenol tetrazolium to determine rapidly MICs of M. tuberculosis to first line anti-mycobacterial drugs. Identification of novel diagnostic tests that can provide a reliable and rapid diagnosis of TB and of drug resistance will contribute substantially to improving TB control globally.
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