ArticleCurrent protocols2025
A Physiologically Relevant In Vitro Model of Nonreplicating Persistent Mycobacterium tuberculosis in Caseum.
Article in Current protocols, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 2 papers.
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Who cites it
2 citing papers in PubMed.
- Mycobacterium tuberculosis long-chain fatty acid resistome reveals universal stress protein TB15.3 as essential for infection.Communications biology · 2026Article
- A Physiologically Relevant In Vitro Model of Nonreplicating Persistent Mycobacterium tuberculosis in Caseum.Current protocols · 2025Article
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Authors and funding
5 authors.
Funding
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Abstract
Tuberculosis (TB) remains one of the leading infectious causes of death worldwide. Persistent bacterial populations in specific microenvironments within the host hamper efficient TB chemotherapy. Caseum in the necrotic core of closed granulomas and cavities of pulmonary TB patients can harbor high burdens of drug-tolerant Mycobacterium tuberculosis (MTB) bacilli, making them particularly difficult to sterilize. Here, we describe protocols for the generation of a surrogate matrix using lipid-rich macrophages to mimic the unique composition of caseum in vivo. Importantly, this caseum surrogate induces metabolic and physiological changes within MTB that reproduce the nonreplicating drug-tolerant phenotype of the pathogen in the native caseous environment, making it advantageous over alternative in vitro models of nonreplicating persistent (NRP) MTB. The protocols include culture of THP-1 monocytes, stimulation of lipid droplet accumulation, lysis and denaturation of the foamy macrophages, inoculation and preadaptation of MTB bacilli in the caseum surrogate, and evaluation of drug bactericidal activity against the NRP population. This novel in vitro model is being used to screen for potent bactericidal antimicrobial agents and to identify vulnerable drug targets, among a variety of other applications, thereby reducing our reliance on in vivo models. © 2025 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Caseum surrogate preparation from γ-irradiated M. tuberculosis-induced foamy THP-1 monocyte-derived macrophages (THPMs) Alternate Protocol 1: Caseum surrogate preparation from stearic acid-induced THPMs Basic Protocol 2: Generation of nonreplicating persistent M. tuberculosis and drug susceptibility testing Alternate Protocol 2: Higher-throughput drug susceptibility screening using caseum surrogate.
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