Early Tuberculosis Immunodetection using Genomic Antigens and Inducement of T-Cells in HIV+TB Subjects in Lagos, Nigeria
A Thesis Submitted to the School of Postgraduate Studies, University of Lagos
The probability of developing tuberculosis (TB) disease is higher in immune compromised subjects such as HIV/AIDS patients. Conventional methods of diagnosing TB which include sputum smear microscopy, culture and chest X-ray have drawbacks and established limitations. The limitations are pronounced in immuno-compromised subjects. The consequence of poor diagnosis of TB especially in resources limited environment like Nigeria contributes to the rising incidence of TB. Introducing immuno detection method using genomic antigens is priming for early detection of TB. This study was designed to evaluate the available genomic TB antigens for T-cell stimulation and to explore possibility of improving diagnostic performance in immuno-compromised patient by inducing T-cells. The study population includes subjects with active TB only, HIV+TB, HIV only, subjects with TB contacts and apparently healthy controls. Smear microscopy and culture were performed on sputum samples obtained. Whole blood cells from suspected TB infected subjects and TB contacts were incubated with RPMI 1640 and glutamine to provide enriched medium of protein and vitamins for the T-cells in the blood and fetal calf serum (FCS) to induce T-cells during incubation so as to make the T-cells respond better when stimulated during incubation. Low dosage of 2.5 µg/ml of TB antigens (ESAT-6,CFP-10 and ELLI the new TB antigen) obtained from LIONEX Inc. Germany was used to stimulate the T-cells in the whole blood during the 72 hour incubation. Sera obtained after stimulation were used for Interferon Gamma Release Assay (IGRA) to assess the IFN-γ released during T-cell stimulation. The result of smear microscopy and culture revealed that even though a high percentage of positivity was obtained from the result of smear microscopy and culture, some of the positives (20.83%) smear microscopy were negative for culture. These were part of the drawbacks of high sensitivity and low specificity observed. Result of using TB antigens for T-cell stimulation showed that ELLI the new antigen was the most immunogenic of the antigens used when compared with existing ESAT-6 and CFP-10. T-cells performed optimally when induced with 1% FCS and stimulated with low dose antigen of ELLI and the combined ESAT-6+CFP-10. The inducement of T-cells improved the diagnosis of TB in HIV subjects that were asymptomatic of TB and individuals at risk especially the previous TB contacts. Sensitivity and specificity of IGRA increased above 80% with the inducement of T-cells. In conclusion, the study attempts to improve TB diagnosis using genomic antigens relative to the conventional non genomic antigens methods. This approach has the potential to improve detection of TB subjects using immunodetection technique.