Rapid and Sensitive Detection of Tuberculosis by MS/MS in Clinical

PRINCIPAL INVESTIGATOR: Sonesson, Anders AWARDEE ORGANIZATION: Microbial Insights, Inc. 201 Center Park Dr., Suite 1140 Knoxville, TN 37922 TELEPHONE NUMBER: (615) 966-7356 AWARDEE NUMBER: 200-93-0681 In response to the resurgent epidemic of tuberculosis which has been greatly stimulated by the large increase in immuno-compromised persons, it is imperative that rapid and accurate diagnostic tests be available. We propose to establish a test system based on lipid constituents found in the waxes of Mycobacteria for detection and species identification. The "signature" lipids are recovered in sputum, tissue specimens, cerebrospinal fluid, etc., to establish the diagnosis. Components of the Mycobacterial waxes have sufficiently different proportions that they can be used to identify the specific species (which has critical therapeutic implications). Two constituents will be measured. One, tuberulostearic acid is found in the Actinomycetales (of which the Mycobacteria are a part) that can be measured at great sensitivity. The other is a series of multimethyl long chain fatty acids found exclusively in the Mycobacteria. Proportions of mycocersatic acids can be utilized to identify the species of Mycobacteria. These lipids can be quantitatively recovered from clinical specimens and identified chemically without the requirements that the bacteria be isolated and cultured. Application of supercritical fluid extraction (potential one-day sample processing), improved purification methods, with detection by tandem mass spectrometry greatly increase the selectivity and sensitivity (possibly to 6-60 organisms) provide a rapid and quantitative assessment of Mycobacterial infection. Tuberculosis is becoming such an important medical problem that performing this test expeditiously is a viable commercial enterprise.