Biology subjects

Gouzy, A.

Publications and source records attributed to Gouzy, A..

4 recordsLinked to original sources

bioRxiv2026

Non-canonical peptidoglycan cross-linking is essential for Mycobacterium tuberculosis acid resistance

Mycobacterium tuberculosis withstands acidic conditions to survive and replicate within macrophages. To define the genetic determinants of this adaptation, we performed a transposon screen in a lipid-rich, acidic medium that mimics the host environment and supports robust M. tuberculosis growth. This screen identified ldtB, encoding an L,D-transpeptidase, as essential for growth and survival under acid stress. Loss of LdtB decreased 3-3 peptidoglycan cross-linking, disrupted cell wall architecture, and impaired intrabacterial pH homeostasis, resulting in increased susceptibility to cell wall-active antibiotics. Notably, M. tuberculosis lacking LdtB displayed heightened sensitivity to meropenem within macrophages, suggesting that targeting this enzyme could potentiate {beta}-lactam efficacy during infection. These findings establish LdtB as a key mediator linking peptidoglycan homeostasis to acid stress resistance and underscores the importance of in vitro culture models that recapitulate the host microenvironment for uncovering new in vivo active therapeutic targets. TeaserDisabling a key peptidoglycan cross-linking enzyme compromises M. tuberculosis survival under acidic stress and antibiotic exposure.

microbiology↗

bioRxiv2026

Metabolic control of drug resistance by a mycobacterial ion channel

Pyrazinamide (PZA) is a cornerstone of modern tuberculosis therapy, yet its context-dependent activity has obscured both its mode of action and resistance mechanisms. Using a host-mimicking culture system integrated with genome-wide CRISPRi profiling, metabolomics, and comparative genomics, we identify a previously unrecognized driver of PZA resistance in humans: loss of the ion channel Rv2571c. Rv2571c mediates -ketoglutarate efflux, amplifying PZA-induced cytoplasmic acidification under host-relevant acidic conditions. Loss-of-function mutations confer resistance in vitro and in vivo and are under positive selection in clinical isolates, establishing this pathway as a resistance determinant in patients. Together, these findings define a novel, ion channel-mediated resistance mechanism, establish cytoplasmic acidification as the basis of PZA killing, and inform resistance detection and treatment-shortening drug development.

microbiology↗

bioRxiv2025

Pyrazinamide kills Mycobacterium tuberculosis via pH-driven weak-acid permeation and cytosolic acidification

Pyrazinamide (PZA) is a cornerstone drug in tuberculosis (TB) treatment with a strong bactericidal activity in vivo on both actively and non-replicating bacterial subpopulations. Yet the precise mode of action of its active form, pyrazinoic acid (POA), remains unclear. In this study, we comprehensively explore and challenge the two major and conflicted models of PZA mode of action. The pH-dependent model, where the drug is mostly effective at acidic pH by acidifying Mycobacterium tuberculosis (Mtb) cytosol, and the PanD-dependent model where PZA active form targets the aspartate decarboxylase PanD, therefore depleting pantothenate (Panto) and subsequently coenzyme A (CoA) levels regardless of the surrounding pH. By combining standard antimicrobial susceptibility testing at various pH with fluorescence-based live recording of Mtb intrabacterial pH (IBpH), we demonstrate that PZA kills Mtb by decreasing IBpH, independently of Panto levels. Comparative studies between a prototrophic Mtb strain and a Panto auxotrophic mutant lacking the panCD locus confirmed that PZA bactericidal activity is primarily driven by pH and its ability to acidify Mtb cytosol, independently of the aspartate decarboxylase PanD. Bio-electrophysiology experiments revealed that acidic pH promotes the conversion of the pyrazinoate anion POA- into HPOA which in turn acts as conventional weak acid that facilitates membrane permeation and cytosolic acidification. Finally, using custom culture media, we demonstrate that PZA displays heterogeneous efficacy according to the media composition, therefore proposing a revisited biological model that might explain the discrepancies around PZA unique mode of action. Overall, this work constitutes the first comprehensive side-by-side investigation of the two models and univocally supports a pH-dependent mechanism of action underlying PZA sterilizing activity, providing new insights for the development of more effective PZA-like drugs.

microbiology↗

bioRxiv2022

Reliable detection of pyrazinamide antitubercular activity in vitro.

Pyrazinamide (PZA) is a pivotal antibiotic for the chemotherapy of tuberculosis (TB), a disease caused by the bacterium Mycobacterium tuberculosis (Mtb). PZA is notorious for its poor in vitro activity which complicates phenotypic PZA-susceptibility testing (PPST) and likely causes inappropriate treatment of TB patients. Here, we show that PZA activity can be reliably detected using an acidic and lipid-rich culture medium mimicking conditions Mtb encounters during an infection. Our growth model could facilitate PPST to improve the treatment of TB patients and ameliorate the global surveillance of PZA resistance.

microbiology↗