title: Assessment of in vitro immunity to Mycobacterium tuberculosis in a human peripheral blood infection model using a luciferase reporter construct of M. tuberculosis H37Rv.
abstract: Protective immune responses to tuberculosis in man are primarily cell-mediated and require the interaction of specific T cells, cytokines and activated macrophages. In the present study, Mycobacterium tuberculosis H37Rv labelled with luciferase reporter enzyme was used to analyse the anti-mycobacterial immunity in man using an in vitro whole blood infection model. Peripheral blood samples obtained from M. bovis bacille Calmette-Guérin (BCG)-vaccinated tuberculin-positive healthy volunteers (n = 23) were cultured with M. tuberculosis H37Rv reporter strain. The growth of bacteria in the whole blood cultures was monitored after 48 and 96 h of infection. The results showed that the growth of M. tuberculosis was significantly inhibited after 96 h (P < 0.029) of culture. Among the cytokines studied, interleukin (IL)-10 and IL-12 were not detected at all, whereas low levels of interferon (IFN)-gamma after 96 h (0.4 IU/ml) and tumour necrosis factor (TNF)-alpha after 48 (135 pg/ml) and 96 h (47 pg/ml) of culture were detected in the supernatants of whole blood infected with M. tuberculosis. The magnitude of bacterial growth correlated directly with the concentration of TNF-alpha detected after 48 h (r = 0.722) and 96 h (r = 0.747) of culture (P <or= 0.0001 and P <or= 0.0001, respectively). However, the addition of monoclonal antibodies specific to TNF-alpha and IFN-gamma to the blood cultures did not alter mycobacterial growth indicating the role of other mechanisms/factors in restricting the growth of M. tuberculosis in whole blood cultures.
authors.affiliation: Department of Microbiology, Faculty of Medicine, Kuwait University, Safat, Kuwait. rj.alattiyah@hsc.edu.kw
authors.forename: R
authors.initials: R
authors.lastname: Al-Attiyah
authors.forename: A
authors.initials: A
authors.lastname: El-Shazly
authors.forename: A S
authors.initials: AS
authors.lastname: Mustafa
mesh_terms: D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001500:BCG Vaccine; D001431:Bacteriological Techniques; D003114:Colony-Forming Units Assay; D016207:Cytokines / Q000276:immunology*; D017930:Genes, Reporter; D006801:Humans; D007371:Interferon-gamma / Q000276:immunology; D008156:Luciferases / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000254:growth & development / Q000276:immunology*; D014376:Tuberculosis / Q000276:immunology*; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D001500:BCG Vaccine; D016207:Cytokines; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma; D008156:Luciferases