Fast-growing intracellular Mycobacterium tuberculosis populations evade antibiotic treatment
Abstract
Slow growth rates are considered a hallmark of Mycobacterium tuberculosis (Mtb) and have been historically associated with persistence and subsequent drug tolerance. Despite the fundamental role intracellular growth plays in the pathogenesis of tuberculosis (TB), approaches that define intracellular growth rates have remained challenging. Here, we developed a highthroughput, live-cell imaging approach to quantify Mtb replication within human macrophage populations at high spatiotemporal, single-cell resolution. Unexpectedly, we discovered fast-growing intracellular Mtb populations with doubling times below 10 hours. Importantly, when treated with first-line antibiotics, these intracellular fast-growing subpopulations remained. To validate this in vivo, we developed a mouse model of tuberculosis featuring a Mtb-Timer fluorophore that reports replication activity. Single-cell analysis revealed a strong correlation between heavily burdened host cells and Mtb populations with heightened replicative activity. These data identify fast-growing intracellular bacteria as a mechanism of antibiotic evasion and challenges the prevailing idea that attributes antibiotic tolerance to bacterial dormancy.Tuberculosis (TB) is one of the most devastating infectious diseases throughout human history, affecting humanity for over 15,000 years 1 and resulting in 1.23 million deaths in 2024 2 . The current chemotherapy regimen for TB requires at least four months and four antibiotics to cure the disease 3 . The reasons behind this prolonged treatment duration and multi-drug requirements are multifactorial and remain poorly understood 4-6 .
Mycobacterium tuberculosis (Mtb), the causative agent of TB, can adopt an intracellular lifestyle and replicate within host cells during infection 7 . In the host, Mtb faces diverse intracellular and extracellular microenvironments that results in single-cell phenotypic variability 8,9 . Compelling evidence indicates a host cell contribution to antibiotic accumulation and efficacy 10-14 , but the underlying mechanisms remain poorly defined.
Mtb is canonically defined as a slow growing bacterium, characterised by bulk population doubling times of approximately 18-24 h 15 . These slow or non-growing bacterial phenotypes are frequently associated with the ability of Mtb to evade antibiotic treatment 16-18 . Previous
Related
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BibTeX
@article{day2026fastgrowing,
title = {{Fast-growing intracellular Mycobacterium tuberculosis populations evade antibiotic treatment}},
author = {Nathan J Day and Baptiste Pradel and Chak Hon Luk and Pierre Santucci and Antony Fearns and Angela Rodgers and Beren Aylan and Laure Botella and Julien Vaubourgeix and Julio Ortiz Canseco and Natalia Athanasiadi and Maximiliano G Gutierrez},
journal = {Nature Communications},
year = {2026},
month = jul,
doi = {10.1038/s41467-026-75915-8},
url = {https://hal.inrae.fr/hal-05748460v1},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Fast-growing intracellular Mycobacterium tuberculosis populations evade antibiotic treatment
AU - Nathan J Day
AU - Baptiste Pradel
AU - Chak Hon Luk
AU - Pierre Santucci
AU - Antony Fearns
AU - Angela Rodgers
AU - Beren Aylan
AU - Laure Botella
AU - Julien Vaubourgeix
AU - Julio Ortiz Canseco
AU - Natalia Athanasiadi
AU - Maximiliano G Gutierrez
PY - 2026
DA - 2026/07/28
JO - Nature Communications
DO - 10.1038/s41467-026-75915-8
UR - https://hal.inrae.fr/hal-05748460v1
AB - Slow growth rates are considered a hallmark of Mycobacterium tuberculosis (Mtb) and have been historically associated with persistence and subsequent drug tolerance. Despite the fundamental role intracellular growth plays in the pathogenesis of tuberculosis (TB), approaches that define intracellular growth rates have remained challenging. Here, we developed a highthroughput, live-cell imaging approach to quantify Mtb replication within human macrophage populations at high spatiotemporal, single-cell resolution. Unexpectedly, we discovered fast-growing intracellular Mtb populations with doubling times below 10 hours. Importantly, when treated with first-line antibiotics, these intracellular fast-growing subpopulations remained. To validate this in vivo, we developed a mouse model of tuberculosis featuring a Mtb-Timer fluorophore that reports replication activity. Single-cell analysis revealed a strong correlation between heavily burdened host cells and Mtb populations with heightened replicative activity. These data identify fast-growing intracellular bacteria as a mechanism of antibiotic evasion and challenges the prevailing idea that attributes antibiotic tolerance to bacterial dormancy.Tuberculosis (TB) is one of the most devastating infectious diseases throughout human history, affecting humanity for over 15,000 years 1 and resulting in 1.23 million deaths in 2024 2 . The current chemotherapy regimen for TB requires at least four months and four antibiotics to cure the disease 3 . The reasons behind this prolonged treatment duration and multi-drug requirements are multifactorial and remain poorly understood 4-6 . Mycobacterium tuberculosis (Mtb), the causative agent of TB, can adopt an intracellular lifestyle and replicate within host cells during infection 7 . In the host, Mtb faces diverse intracellular and extracellular microenvironments that results in single-cell phenotypic variability 8,9 . Compelling evidence indicates a host cell contribution to antibiotic accumulation and efficacy 10-14 , but the underlying mechanisms remain poorly defined. Mtb is canonically defined as a slow growing bacterium, characterised by bulk population doubling times of approximately 18-24 h 15 . These slow or non-growing bacterial phenotypes are frequently associated with the ability of Mtb to evade antibiotic treatment 16-18 . Previous
SN - 2041-1723
SN - 2041-1723
ER -