Structure–function relationships of the antigenicity of mycolic acids in tuberculosis patients

Mervyn Beukes,Yolandy Lemmer,Madrey Deysel,Juma’A R Al Dulayymi,Mark S Baird,Gani Koza,Maximiliano M Iglesias,Richard R Rowles,Cornelia Theunissen,Johan Grooten,Gianna Toschi,Vanessa V Roberts,Lynne Pilcher,Sandra Van Wyngaardt,Nsovo Mathebula,Mohammed Balogun,Anton C Stoltz,Jan A Verschoor

doi:10.1016/j.chemphyslip.2010.09.006

Mervyn Beukes, Yolandy Lemmer + Show 16 more

Open Access

https://doi.org/10.1016/j.chemphyslip.2010.09.006

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Abstract

Cell wall mycolic acids (MA) from Mycobacterium tuberculosis (M.tb) are CD1b presented antigens that can be used to detect antibodies as surrogate markers of active TB, even in HIV coinfected patients. The use of the complex mixtures of natural MA is complicated by an apparent antibody cross-reactivity with cholesterol. Here firstly we report three recombinant monoclonal scFv antibody fragments in the chicken germ-line antibody repertoire, which demonstrate the possibilities for cross-reactivity: the first recognized both cholesterol and mycolic acids, the second mycolic acids but not cholesterol, and the third cholesterol but not mycolic acids. Secondly, MA structure is experimentally interrogated to try to understand the cross-reactivity. Unique synthetic mycolic acids representative of the three main functional classes show varying antigenicity against human TB patient sera, depending on the functional groups present and on their stereochemistry. Oxygenated (methoxy- and keto-) mycolic acid was found to be more antigenic than alpha-mycolic acids. Synthetic methoxy-mycolic acids were the most antigenic, one containing a trans-cyclopropane apparently being somewhat more antigenic than the natural mixture. Trans-cyclopropane-containing keto- and hydroxy-mycolic acids were also found to be the most antigenic among each of these classes. However, none of the individual synthetic mycolic acids significantly and reproducibly distinguished the pooled serum of TB positive patients from that of TB negative patients better than the natural mixture of MA. This argues against the potential to improve the specificity of serodiagnosis of TB with a defined single synthetic mycolic acid antigen from this set, although sensitivity may be facilitated by using a synthetic methoxy-mycolic acid.

Highlights

South Africa currently has the highest per capita incidence of TB in the world
The mycolic acids (MA) methyl-ester is not recognized by antibodies, whereas the free MA (MA mix) is recognized by both TB pos and TB neg patient sera, but more strongly with TB pos sera. This supports the hypothesis that the enzyme-linked immunosorbent assay (ELISA) antibody binding signal is due to recognition of an antigen consisting of one or more MAs, in which the hydroxyl group of the free MA-carboxylic acid probably participates in interor intramolecular stabilization of a specific antigen conformation
Do they act as pathogen associated molecular patterns (PAMP) for induction of murine innate immunity (Korf et al, 2005), but they are able to reprogramme murine macrophages to modulate their inflammatory activity (Korf et al, 2006)

Summary

Introduction

South Africa currently has the highest per capita incidence of TB in the world. In 2007 alone 112,000 people died of TB in South Africa, of whom 94,000 were co-infected with HIV (WHO, 2009). One of the biggest challenges facing clinicians is the time it takes to accurately diagnose TB. Using the conventional methods, it takes on average 4 weeks to diagnose TB, which leads to a delay in treatment of the disease. Two thirds of TB deaths could be prevented by early diagnosis.

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