New synthetic lipid antigens for rapid serological diagnosis of tuberculosis

Abstract

BackgroundDuring pulmonary tuberculosis (PTB) antibodies are generated to trehalose esters of mycolic acids which are cell wall lipids of Mycobacterium tuberculosis (Mtb). Attempts have been made to use these complex natural mixtures in serological tests for PTB diagnosis.AimThe aim of this work was to determine whether a serological test based on a panel of defined individual trehalose esters of characteristic synthetic mycolic acids has improved diagnostic accuracy in distinguishing patients with culture positive PTB from individuals who were Mtb culture negative.MethodOne hundred serum samples from well-characterized patients with presumptive tuberculosis, and diagnosed as having pulmonary smear and culture positive TB, or being culture and smear negative were evaluated by ELISA using different combinations of synthetic antigens and secondary antibodies. Using cut-off values determined from these samples, we validated this study blind in samples from a further 249 presumptive TB patients.ResultsWith the first 100 samples, detailed responses depended both on the precise structure of the antigen and on the secondary antibody. Using a single antigen, a sensitivity/specificity combination for smear and culture positive PTB detection of 85 and 88% respectively was achieved; this increased to 96% and 95% respectively by a statistical combination of the results with seven antigens. In the blind study a sensitivity/specificity of 87% and 83% was reached with a single antigen. With some synthetic antigens, the responses from all 349 samples were significantly better than those with the natural mixture. Combining the results for seven antigens allowed a distinction between culture positive and negative with a ROC AUC of 0.95.ConclusionWe have identified promising antigen candidates for serological assays that could be used to diagnose PTB and which could be the basis of a much-needed, simple, rapid diagnostic test that would bring care closer to communities.

Update on tuberculosis biomarkers: From correlates of risk, to correlates of active disease and of cure from disease

“…Recently, an assay based on antibodies to the trehalose esters of mycolic acids, which are Mtb‐specific cell wall glycolipids, was reported. This assay showed promising accuracy with a sensitivity and specificity of 87% and 83%, respectively …”

“…This assay showed promising accuracy with a sensitivity and specificity of 87% and 83%, respectively. 75 Interestingly, a multiplex TB serology panel using microbead suspension arrays containing a combination of 11 Mtb antigens was shown to have an overall sensitivity of 91% in serum/plasma samples from cultureproven TB patients. Test specificity was 96% compared to chronic obstructive pulmonary disease patients and 91% compared to the healthy population.…”

Revealing solvent-dependent folding behavior of mycolic acids from Mycobacterium tuberculosis by advanced simulation analysis

“…Here, two key features play a significant role; oxygenation of the MA [15][16][17][18][19][20][21][22][23], and cyclopropanation [24][25][26][27], including the specific stereochemistry of the cyclopropane ring [22][23][24][25]. Confirmation of the role of individual MA structures and benchmarked mixtures has recently been achieved through the use of synthetic derivatives [20][21][22][23][31][32][33], and strongly implies that the underlying MA structure (individually or as sugar-ester derivatives), which steers the physical properties and conformational behavior of the molecule, can offer a window to rationalize their biological role.…”

“…MAs are central to the host immune response against the organism and have been shown to be good antigens for use in serodiagnosis of TB [30,31,[33][34][35][36]. However, fundamental details such as how MAs are arranged in the cell wall, and how they interact with immune components, are yet to be determined.…”

The synthesis of single enantiomers of trans-alkene containing mycolic acids and related sugar esters

“…63 Such sugar esters are also antigenic to antibodies in serum of patients infected with active TB. 64 The trans-alkene sugar esters described above were each evaluated as antigens in ELISA for responses to antibodies in serum of patients infected with pulmonary tuberculosis, using modified IgG(Fc) as secondary antibody to provide a colour response. Twenty samples from a WHO Specimen Bank were used, 9 culture and smear positive for TB, 11 smear and culture negative and diagnosed as not having TB.…”