Activity of a three-phage combination against Mycobacterium tuberculosis in disease-relevant conditions

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ID: 332520
2026
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Abstract
Abstract Background Phage therapy offers promise to combat antimicrobial resistance, including drug-resistant tuberculosis. Understanding phage activity against Mycobacterium tuberculosis (Mtb) in granuloma-like microenvironments that induce non-replicating states, characterized by hypoxia and acidity, is essential. Objectives To evaluate a phage combination against Mtb under hypoxic, acidic (pH 5.5), and stationary-phase conditions in vitro. Methods Mtb H37Rv cultures were grown to exponential phase for time-kill studies (106 colony forming units (CFU)/mL) under normal, hypoxic, and acidic conditions. Hypoxia was generated using sealed containers with gas sachets. Stationary-phase cultures were grown for 14–21 days and used undiluted. Treatments included control media, single phages (Fred313cpm-1Δ33, FionnbharthΔ45Δ47and Muddy HRM N0157-2), phage combination (multiplicity of infection 10:1), and rifampicin or isoniazid (0.1 µg/mL each). Phage titers (measured using M. smegmatis overlays) and bacterial counts (CFU on Middlebrook 7H11) were assessed at repeated time-points. Longitudinal phage-bacteria dynamics mixed-effects modelling and simulation was performed. Results In actively growing cultures, phage levels rose by day 7, followed by sustained reductions in Mtb counts over 31 days. Phages prevented regrowth seen with antibiotics alone. Acidic conditions variably affected individual phage stability and activity. Under hypoxic and stationary conditions, phage titers remained stable, but no reduction in Mtb was observed. Model-based predictions adequately captured phage-mycobacterial interactions with and without rifampicin. Conclusions Productive lytic activity was not observed in assays with non-replicating mycobacteria. Established models can support future experimental designs. This study provides mechanistic insights driving progress in the field of phage therapy development.
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Authors Saskia Janssen, S E Larsen, M Pozuelo Torres, Massinissa Beldjenna, Carlos A. Guerrero-Bustamante, I Florian, T Smytheman, T Guo, R C van Wijk, G F Hatfull, A H Diacon, R N Coler, Jakko van Ingen
Journal The Journal of infectious diseases
Year 2026
DOI
10.1093/infdis/jiag516
URL
https://doi.org/10.1093/infdis/jiag516
Keywords Keywords not found

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