Implementing best practices on data generation and reporting of Mycobacterium tuberculosis in vitro assays within the ERA4TB consortium

Wijk, Rob C. van; Lucía, Ainhoa; Sudhakar, Pavan Kumar; Sonnenkalb, Lindsay; Gaudin, Cyril; Hoffmann, Eik; Dremierre, Bérénice; Aguilar-Ayala, Diana Angélica; Molin, Michael Dal; Rybniker, Jan; Giorgi, Stefano De; Cioetto-Mazzabò, Laura; Segafreddo, Greta; Manganelli, Riccardo; Degiacomi, Giulia; Recchia, Deborah; Pasca, Maria Rosalia; Simonsson, Ulrika S. H.; Ramón-García, Santiago

doi:10.1016/j.isci.2023.106411

Cited by 2 publications

(1 citation statement)

References 24 publications

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“…Bacterial viability in the cultures exposed to stress and used to collect RNA did not show any statistically significant variation during the entire course of the experiment ( van Wijk et al, 2023 ) ( Supplementary Figure S1 ). The bacterial strains used in this work are listed in Table 1 .…”

Section: Methodsmentioning

confidence: 99%

Structure of the SigE regulatory network in Mycobacterium tuberculosis

Cioetto-Mazzabò,

Sorze,

Babic

et al. 2024

Front. Microbiol.

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SigE is one of the main regulators of mycobacterial stress response and is characterized by a complex regulatory network based on two pathways, which have been partially characterized in conditions of surface stress. The first pathway is based on the induction of sigE transcription by the two-component system MprAB, while the second is based on the degradation of SigE anti-sigma factor RseA by ClpC1P2, a protease whose structural genes are induced by ClgR. We characterized the dynamics of the SigE network activation in conditions of surface stress and low pH in Mycobacterium tuberculosis. Using a series of mutants in which the main regulatory nodes of the network have been inactivated, we could explore their hierarchy, and we determined that MprAB had a key role in the network activation in both stress conditions through the induction of sigE. However, while in conditions of surface stress the absence of MprAB totally abrogated sigE induction, under low pH conditions it only resulted in a small delay of the induction of sigE. In this case, sigE induction was due to SigH, which acted as a MprAB backup system. The ClgR pathway, leading to the degradation of the SigE anti-sigma factor RseA, was shown to be essential for the activation of the SigE network only following surface stress, where it showed an equal hierarchy with the MprAB pathway.

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Section: Methodsmentioning

confidence: 99%

Structure of the SigE regulatory network in Mycobacterium tuberculosis

Cioetto-Mazzabò,

Sorze,

Babic

et al. 2024

Front. Microbiol.

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Evaluation of critical parameters in the hollow‐fibre system for tuberculosis: A case study of moxifloxacin

Aguilar‐Ayala,

Sanz‐García,

Rabodoarivelo

et al. 2024

Brit J Clinical Pharma

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AimsThe hollow‐fibre system for tuberculosis (HFS‐TB) is a preclinical model qualified by the European Medicines Agency to underpin the anti‐TB drug development process. It can mimic in vivo pharmacokinetic (PK)–pharmacodynamic (PD) attributes of selected antimicrobials, which could feed into in silico models to inform the design of clinical trials. However, historical data and published protocols are insufficient and omit key information to allow experiments to be reproducible. Therefore, in this work, we aim to optimize and standardize various HFS‐TB operational procedures.MethodsFirst, we characterized bacterial growth dynamics with different types of hollow‐fibre cartridges, Mycobacterium tuberculosis strains and media. Second, we mimicked a moxifloxacin PK profile within hollow‐fibre cartridges, in order to check drug–fibres compatibility. Lastly, we mimicked the moxifloxacin total plasma PK profile in human after once daily oral dose of 400 mg to assess PK–PD after different sampling methods, strains, cartridge size and bacterial adaptation periods before drug infusion into the system.ResultsWe found that final bacterial load inside the HFS‐TB was contingent on the studied variables. Besides, we demonstrated that drug–fibres compatibility tests are critical preliminary HFS‐TB assays, which need to be properly reported. Lastly, we uncovered that the sampling method and bacterial adaptation period before drug infusion significantly impact actual experimental conclusions.ConclusionOur data contribute to the necessary standardization of HFS‐TB experiments, draw attention to multiple aspects of this preclinical model that should be considered when reporting novel results and warn about critical parameters in the HFS‐TB currently overlooked.

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Implementing best practices on data generation and reporting of Mycobacterium tuberculosis in vitro assays within the ERA4TB consortium

Cited by 2 publications

References 24 publications

Structure of the SigE regulatory network in Mycobacterium tuberculosis

Structure of the SigE regulatory network in Mycobacterium tuberculosis

Evaluation of critical parameters in the hollow‐fibre system for tuberculosis: A case study of moxifloxacin

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