Assessment of in vitro immunity to Mycobacterium tuberculosis in a human peripheral blood infection model using a luciferase reporter construct of M. tuberculosis H37Rv

Summary 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)‐γ after 96 h (0·4 IU/ml) and tumour necrosis factor (TNF)‐α 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‐α detected after 48 h ( r  = 0·722) and 96 h ( r  = 0·747) of culture ( P  ≤ 0·0001 and P  ≤ 0·0001, respectively). However, the addition of monoclonal antibodies specific to TNF‐α and IFN‐γ 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.