Browsing by Author "Jason Wagner"

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    Serological diagnosis of pulmonary Mycobacterium tuberculosis infection by LIPS using a multiple antigen mixture
    (2015-10-08) Peter D. Burbelo; Jason Keller; Jason Wagner; James S. Klimavicz; Ahmad Bayat; Craig S. Rhodes; Bassirou Diarra; Ploenchan Chetchotisakd; Yupin Suputtamongkol; Sasisopin Kiertiburanakul; Steven M. Holland; Sarah K. Browne; Sophia Siddiqui; Joseph A. Kovacs; National Institute of Dental and Craniofacial Research; University of Sciences; Khon Kaen University; Mahidol University; National Institute of Allergy and Infectious Diseases; NIH Clinical Center
    © 2015 Burbelo et al. Background: There is an urgent need for a simple and accurate test for the diagnosis of human Mycobacterium tuberculosis, the infectious agent causing tuberculosis (TB). Here we describe a serological test based on light emitting recombinant proteins for the diagnosis of pulmonary Mycobacterium tuberculosis infection. Methods: Luciferase Immunoprecipitation Systems (LIPS), a fluid-phase immunoassay, was used to examine antibody responses against a panel of 24 different M. tuberculosis proteins. Three different strategies were used for generating the constructs expressing the recombinant fusion M. tuberculosis proteins with luciferase: synthetic gene synthesis, Gateway recombination cloning, and custom PCR synthesis. A pilot cohort of African pulmonary TB patients was used for initial antibody screening and confirmatory studies with selected antigens were performed with a cohort from Thailand and healthy US blood donors. In addition to testing M. tuberculosis antigens separately, a mixture that tested seven antigens simultaneously was evaluated for diagnostic performance. Results: LIPS testing of a pilot set of serum samples from African pulmonary TB patients identified a potential subset of diagnostically useful M. tuberculosis antigens. Evaluation of a second independent cohort from Thailand validated highly significant antibody responses against seven antigens (PstS1, Rv0831c, FbpA, EspB, bfrB, HspX and ssb), which often showed robust antibody levels up to 50- to 1000-fold higher than local community controls. Marked heterogeneity of antibody responses was observed in the patients and the combined results demonstrated 73.5 % sensitivity and 100 % specificity for detection of pulmonary TB. A LIPS test simultaneously employing the seven M. tuberculosis antigen as a mixture matched the combined diagnostic performance of the separate tests, but showed an even higher diagnostic sensitivity (90 %) when a cut-off based on healthy US blood donors was used. Conclusion: A LIPS immunoassay employing multiple M. tuberculosis antigens shows promise for the rapid and quantitative serological detection of pulmonary TB.