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Original Research ARTICLE Provisionally accepted The full-text will be published soon. Notify me

Front. Microbiol. | doi: 10.3389/fmicb.2019.01922

Comprehensive characterization of the attenuated double auxotroph Mycobacterium tuberculosis ΔleuDΔpanCD as an alternative to H37Rv

  • 1Centre for Tuberculosis Research, South African Medical Research Council, South Africa
  • 2Faculty of Medicine and Health Sciences, Stellenbosch University, South Africa
  • 3Institute for Cell and Molecular Biosciences, Newcastle University, United Kingdom
  • 4Amsterdam Institute for Molecules, Medicines and Systems, VU University Amsterdam, Netherlands

Currently available model organisms such as Mycobacterium smegmatis and Mycobacterium bovis Bacillus Calmette-Guérin (BCG) have significantly contributed to our understanding of tuberculosis (TB) biology, these models have limitations such as differences in genome size, growth rates and virulence. However, attenuated Mycobacterium tuberculosis strains may provide more representative, safer models to study M. tuberculosis biology. For example, the M. tuberculosis ΔleuDΔpanCD double auxotroph, has undergone rigorous in vitro and in vivo safety testing. Like other auxotrophic strains, this has subsequently been approved for use in biosafety level (BSL) 2 facilities. Auxotrophic strains have been assessed as models for drug-resistant M. tuberculosis and for studying latent TB. These offer the potential as safe and useful models, but it is important to understand how well these recapitulate salient features of non-attenuated M. tuberculosis. We therefore performed a comprehensive comparison of M. tuberculosis H37Rv and M. tuberculosis ΔleuDΔpanCD. These strains demonstrated similar in vitro and intra-macrophage replication rates, similar responses to anti-TB agents and whole genome sequence conservation. Shotgun proteomics analysis suggested that M. tuberculosis ΔleuDΔpanCD has a heightened stress response that leads to reduced bacterial replication during exposure to acid stress, which has been verified using a dual-fluorescent replication reporter assay. Importantly, infection of human peripheral blood mononuclear cells with the 2 strains elicited comparable cytokine production, demonstrating the suitability of M. tuberculosis ΔleuDΔpanCD for immunological assays. We provide comprehensive evidence to support the judicious use of M. tuberculosis ΔleuDΔpanCD as a safe and suitable model organism for M. tuberculosis research, without the need for a BSL3 facility.

Keywords: biosafety level 2, attenuated auxotroph, model organism, Mycobacterium tuberculosis, H37Rv

Received: 15 May 2019; Accepted: 05 Aug 2019.

Edited by:

Mattias Collin, Lund University, Sweden

Reviewed by:

Mario Alberto Flores-Valdez, CONACYT Centro de Investigación y Asistencia en Tecnología y Diseño del Estado de Jalisco (CIATEJ), Mexico
FABIANA BIGI, National Institute of Agricultural Technology, Argentina  

Copyright: © 2019 Mouton, Heunis, Dippenaar, Gallant, Kleynhans and Sampson. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Prof. Samantha L. Sampson, Centre for Tuberculosis Research, South African Medical Research Council, Cape Town, South Africa, ssampson@sun.ac.za