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A High-throughput Compatible Assay to Evaluate Drug Efficacy against Macrophage Passaged Mycobacterium tuberculosis
Published on: March 24, 2017
MspA-Mycobacterium tuberculosis-transformant with reduced virulence: the "unbirthday paradigm"
Otmane Lamrabet1, Eric Ghigo1, Jean-Louis Mège1
1Aix Marseille Université, URMITE, UM63, CNRS UMR7278, IRD 198, Inserm 1095, 13005 Marseille, France.
Abstract:
Expressing mspA porin gene from Mycobacterium smegmatis in Mycobacterium tuberculosis attenuated this pathogen. Intracellular growth of the transformants into free-living amoeba and murine and human macrophages decreased. Furthermore, transformants decreased the microbicidal program of human monocyte-derived macrophages. BALB/c mice inoculated with transformants exhibited higher weights, lower histological lesions and lower M. tuberculosis inoculum in the liver, spleen and lungs than control mice challenged with wild-type M. tuberculosis. Preliminary evaluation indicated that mice inoculated with this transformant showed higher weights and lower numbers of lung nodules and tissular mycobacteria than control mice when challenged with wild-type M. tuberculosis. Similar to the paradoxical "unbirthday" gift coined by Lewis Carroll in Alice's Adventures in Wonderland, adding mspA gene reduced the virulence of M. tuberculosis and yielded a protective effect. Lost of non-virulence genes is a mechanism for virulence in mycobacteria. Engineering non-virulence genes in M. tuberculosis may yield strains with decreased virulence and increased immunogenicity.
Insights
Introducing the Mycobacterium smegmatis porin A (mspA) gene into Mycobacterium tuberculosis reduced its virulence. This genetic modification attenuated the pathogen, offering a protective effect against tuberculosis in preclinical models.
Area of Science:
- Microbiology
- Immunology
- Genetics
Background:
- Mycobacterium tuberculosis (M. tuberculosis) is a significant human pathogen responsible for tuberculosis.
- Understanding genetic factors that influence M. tuberculosis virulence is crucial for developing novel control strategies.
- The role of porin genes in mycobacterial pathogenesis is an area of ongoing research.
Purpose of the Study:
- To investigate the effect of expressing the mspA porin gene from Mycobacterium smegmatis in M. tuberculosis.
- To assess the impact of mspA gene expression on the virulence and intracellular survival of M. tuberculosis.
- To evaluate the protective potential of M. tuberculosis engineered with the mspA gene in a mouse model.
Main Methods:
- Genetic engineering of M. tuberculosis to express the mspA gene from M. smegmatis.
- In vitro assessment of intracellular growth in amoeba and macrophages (murine and human).
- Evaluation of the microbicidal program in human monocyte-derived macrophages.
- In vivo studies using BALB/c mice challenged with transformant and wild-type M. tuberculosis.
Main Results:
- Expression of the mspA gene significantly attenuated M. tuberculosis.
- Intracellular growth in amoeba and macrophages was reduced in transformant strains.
- The microbicidal program of human macrophages was less suppressed by transformants.
- Mice infected with transformants showed improved weight gain, reduced tissue lesions, and lower bacterial loads in organs compared to controls.
- Preliminary data suggested reduced lung nodules and tissue mycobacteria in mice challenged with wild-type M. tuberculosis after prior inoculation with the transformant.
Conclusions:
- The mspA gene from M. smegmatis can attenuate the virulence of M. tuberculosis.
- Engineering M. tuberculosis with non-virulence genes like mspA may lead to reduced pathogenicity.
- This approach holds potential for developing live attenuated vaccines or therapeutic strategies against tuberculosis.
