Mycobacterium tuberculosis H37Rv comparative gene-expression analysis in synthetic medium and human macrophage

Mycobacteria are intracellular pathogens that survive and grow in host macr ophages. Following phagocytosis, sustained intracellular bacterial growth d epends on its ability to avoid destruction by macrophage-mediated host defe nces such as lysosomal enzymes, reactive oxygen and the reactive nitrogen i ntermediates. This suggests that the interaction between host cell and microbe is delicat ely balanced, and can be tipped in favour of either organism. The identific ation of Mycobacterium tuberculosis H37Rv (MTB) genes expressed within host cells would contribute greatly to the development of new strategies to fig ht tuberculosis. In the present study, we compared MTB gene expression in t he course of intra- ( human macrophages) and extracellular growth (Sauton's medium) to ascertain whether differences might occur between gene-expressi on patterns in the two habitats of replication. Using reverse-transcriptase polymerase chain reaction (RT-PCR) on a group of 14 MTB-Complex-specific g enes, we found that MT10Sa (a small stable RNA), 35 kDa (unknown), ahpC (al kyl hydroperoxide reductase, AhpC), sigF (alternative RNA Polymerase sigma factor), and katG (catalase-peroxidase, HPI) genes are expressed in both th e environments, while Ag85B, Ag85C (members of the Antigen 85 Complex): rpo V (RNA Polymerase sigma factor) and ESAT6 (early secretory antigen, 6 kDa) are expressed only in the in vitro culture; on the other hand, Ag85A (Antig en 85 Complex), rpoB (RNA Polymerase beta sub-unit), pab (Protein antigen b ), invA and invB genes (encoding proteins that show homologies with p60 of Listeria monocytogenes) are expressed only inside the macrophage. Positive RT-PCR products on cDNAs for these genomic regions were not obtained from a pproximately 1000-fold more bacteria grown in Laboratory Broth. Identificat ion of dl. tuberculosis genes expressed in response to phagocytosis by huma n macrophages increases our basic understanding of the host-pathogen intera ction, and helps to identify bacterial factors necessary for in vivo surviv al and growth. (C) 2000 Elsevier Science B.V, All rights reserved.