Screening strategies for antituberculosis compounds using Mycobacterium tuberculosis are time consuming and require biosafety level 3 (BSL3) facilities, which makes the development of high-throughput assays difficult and expensive. Mycobacterium marinum, a close genetic relative of M. tuberculosis, possesses several advantages as a suitable model for tuberculosis drug screening. However, despite the high genetic similarity, there are some obvious differences in susceptibility to some tuberculosis drugs between these two species, especially for the prodrugs ethionamide and isoniazid. In this study, we aimed to improve M. marinum as a model for antituberculosis drug identification by heterologous expression of two common drug activators, EthA and KatG. These two activators were overexpressed in M. marinum, and the strains were tested against ethionamide, isoniazid, and a library of established antimycobacterial compounds from TB Alliance to compare drug susceptibility. Both in vitro and in vivo using zebrafish larvae, these genetically modified M. marinum strains showed significantly higher susceptibility against ethionamide and isoniazid, which require activation by EthA and KatG. More importantly, a strain overexpressing both ethA and katG was potentially more susceptible to approximately 20% of the antituberculosis hit compounds from the TB Alliance library. Most of these compounds were activated by EthA in M. marinum. Four of these compounds were selected for further analysis, and three of them showed obvious EthA-dependent activity against M. tuberculosis. Overall, our developed M. marinum strains are valuable tools for high-throughput discovery of potential novel antituberculosis prodrugs.
Bibliographical noteFunding Information:
This work was supported from the Netherlands Organization for Scientific Research (NWO) through TTW-NACTAR-16445 granted to W.B. and a VENI grant (016.Veni.171.090) awarded to A.S. The organization Amsterdam Infection and Immunity (AI&II) supported this work with funding awarded to A.S.
© 2021 American Society for Microbiology. All rights reserved.
Copyright 2021 Elsevier B.V., All rights reserved.
- Animal models
- Antimicrobial activity
- Antimicrobial agents
- Enzyme activation
- Mycobacterium tuberculosis