Development of infection model for studying intracellular gene expression of Mycobacterium tuberculosis

Abstract

Summary Mycobacterium tuberculosis complex owe their ability to cause infection because of their intracellular survival ability in professional phagocytic cells of human and the ability to enter into stage of dormancy. The aim of this study was to develop an infection model that could be used to study M. tuberculosis and macrophage interactions at molecular level. Four infection models were examined namely opsonised M bovis BCG / J774.2 macrophage cell line, non-opsonised M. bovis BCG / J774.2 macrophage cell line, opsonised M. tuberculosis I ill4.2 macrophage cell line, and non-opsonised M tuberculosis I ill A.2 macrophage cell line infection models. A J774.2 macrophage cell line was synchronously infected with M. bovis (BCG strain) and M. tuberculosis (H37Rv), respectively at different multiplicity of infections (M.O.I). For opsonisation, the organisms were pre-incubated with human serum prior to infection. The infected cell lines were examined by light microscopy and electron microscopy with viable bacterial counts. Macrophage viability was assessed by trypan blue exclusion staining. The results showed higher significant level of infection of J774.2 macrophage cell line by opsonised M. bovis BCG (30 - 40%) compared to non-opsonised M. bovis BCG (<0.1%) at an M.O.I of 50 (p < 0.05) with high macrophage viability. In contrast, there was no significant statistical difference (p > 0.05) in high infectivity (30 - 42%) with high macrophage viability achieved with using non-opsonised M. tuberculosis and opsonised M. tuberculosis, respectively, at an M.O.I of 10. In conclusion, opsonisation is not required forM tuberculosis I ill42 infection model in contrast to M. bovis BCG / J774.2 infection model where opsonisation is necessary to achieve high level of infection.