Serological diagnosis of pulmonary Mycobacterium tuberculosis infection by LIPS using a multiple antigen mixture.

Abstract

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.

• Authors: 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
• Journal: BMC microbiology (BMC Microbiol) Vol. 15 Pg. 205 (Oct 08 2015) ISSN: 1471-2180 [Electronic] England
• PMID: 26449888 (Publication Type: Evaluation Study, Journal Article, Research Support, N.I.H., Intramural)

Chemical References

• Antibodies, Bacterial
• Antigens, Bacterial
• Luciferases

Topics

• Africa
• Antibodies, Bacterial (blood)
• Antigens, Bacterial (immunology)
• Cohort Studies
• Humans
• Immunoprecipitation (methods)
• Luciferases (analysis)
• Mycobacterium tuberculosis (immunology)
• Pilot Projects
• Sensitivity and Specificity
• Serologic Tests (methods)
• Thailand
• Tuberculosis, Pulmonary (diagnosis)
• United States