Intranasal Administration of Bedaquiline-Loaded Fucosylated Liposomes Provides Anti-Tubercular Activity while Reducing the Potential for Systemic Side Effects.

Liposomal formulations of antibiotics for inhalation offer the potential for the delivery of high drug doses, controlled drug release kinetics in the lung, and an excellent safety profile. In this study, we evaluated the in vivo performance of a liposomal formulation for the poorly soluble, antituberculosis agent, bedaquiline. Bedaquiline was encapsulated within monodisperse liposomes of ∼70 nm at a relatively high drug concentration (∼3.6 mg/mL). Formulations with or without fucose residues, which bind to C-type lectin receptors and mediate a preferential binding to macrophage mannose receptor, were prepared, and efficacy was assessed in an in vivo C3HeB/FeJ mouse model of tuberculosis infection (H37Rv strain). Seven intranasal instillations of 5 mg/kg bedaquiline formulations administered every second day resulted in a significant reduction in lung burden (∼0.4-0.6 Δlog10 CFU), although no differences between fucosylated and nonfucosylated formulations were observed. A pharmacokinetic study in healthy, noninfected Balb/c mice demonstrated that intranasal administration of a single dose of 2.5 mg/kg bedaquiline liposomal formulation (fucosylated) improved the lung bioavailability 6-fold compared to intravenous administration of the same formulation at the same dose. Importantly, intranasal administration reduced systemic concentrations of the primary metabolite, N-desmethyl-bedaquiline (M2), compared with both intravenous and oral administration. This is a clinically relevant finding as the M2 metabolite is associated with a higher risk of QT-prolongation in predisposed patients. The results clearly demonstrate that a bedaquiline liposomal inhalation suspension may show enhanced antitubercular activity in the lung while reducing systemic side effects, thus meriting further nonclinical investigation.

The lipidomics reporting checklist a framework for transparency of lipidomic experiments and repurposing resource data.

The rapid increase in lipidomic studies has led to a collaborative effort within the community to establish standards and criteria for producing, documenting, and disseminating data. Creating a dynamic easy-to-use checklist that condenses key information about lipidomic experiments into common terminology will enhance the field's consistency, comparability, and repeatability. Here, we describe the structure and rationale of the established Lipidomics Minimal Reporting Checklist to increase transparency in lipidomics research.

Lipidation of pneumococcal proteins enables activation of human antigen-presenting cells and initiation of an adaptive immune response.

Streptococcus pneumoniae remains a significant global threat, with existing vaccines having important limitations such as restricted serotype coverage and high manufacturing costs. Pneumococcal lipoproteins are emerging as promising vaccine candidates due to their surface exposure and conservation across various serotypes. While prior studies have explored their potential in mice, data in a human context and insights into the impact of the lipid moiety remain limited. In the present study, we examined the immunogenicity of two pneumococcal lipoproteins, DacB and MetQ, both in lipidated and non-lipidated versions, by stimulation of primary human immune cells. Immune responses were assessed by the expression of common surface markers for activation and maturation as well as cytokines released into the supernatant. Our findings indicate that in the case of MetQ lipidation was crucial for activation of human antigen-presenting cells such as dendritic cells and macrophages, while non-lipidated DacB demonstrated an intrinsic potential to induce an innate immune response. Nevertheless, immune responses to both proteins were enhanced by lipidation. Interestingly, following stimulation of dendritic cells with DacB, LipDacB and LipMetQ, cytokine levels of IL-6 and IL-23 were significantly increased, which are implicated in triggering potentially important Th17 cell responses. Furthermore, LipDacB and LipMetQ were able to induce proliferation of CD4+ T cells indicating their potential to induce an adaptive immune response. These findings contribute valuable insights into the immunogenic properties of pneumococcal lipoproteins, emphasizing their potential role in vaccine development against pneumococcal infections.

LipidSpace: Simple Exploration, Reanalysis, and Quality Control of Large-Scale Lipidomics Studies.

Lipid analysis gained significant importance due to the enormous range of lipid functions, e.g., energy storage, signaling, or structural components. Whole lipidomes can be quantitatively studied in-depth thanks to recent analytical advancements. However, the systematic comparison of thousands of distinct lipidomes remains challenging. We introduce LipidSpace, a standalone tool for analyzing lipidomes by assessing their structural and quantitative differences. A graph-based comparison of lipid structures is the basis for calculating structural space models and subsequently computing lipidome similarities. When adding study variables such as body weight or health condition, LipidSpace can determine lipid subsets across all lipidomes that describe these study variables well by utilizing machine-learning approaches. The user-friendly GUI offers four built-in tutorials and interactive visual interfaces with pdf export. Many supported data formats allow an efficient (re)analysis of data sets from different sources. An integrated interactive workflow guides the user through the quality control steps. We used this suite to reanalyze and combine already published data sets (e.g., one with about 2500 samples and 576 lipids in one run) and made additional discoveries to the published conclusions with the potential to fill gaps in the current lipid biology understanding. LipidSpace is available for Windows or Linux (https://lifs-tools.org).

Global analysis of putative phospholipases in Plasmodium falciparum reveals an essential role of the phosphoinositide-specific phospholipase C in parasite maturation.

For its replication within red blood cells, the malaria parasite depends on a highly active and regulated lipid metabolism. Enzymes involved in lipid metabolic processes such as phospholipases are, therefore, potential drug targets. Here, using reverse genetics approaches, we show that only 1 out of the 19 putative phospholipases expressed in asexual blood stages of Plasmodium falciparum is essential for proliferation in vitro, pointing toward a high level of redundancy among members of this enzyme family. Using conditional mislocalization and gene disruption techniques, we show that this essential phosphoinositide-specific phospholipase C (PI-PLC, PF3D7_1013500) has a previously unrecognized essential role during intracellular parasite maturation, long before its previously perceived role in parasite egress and invasion. Subsequent lipidomic analysis suggests that PI-PLC mediates cleavage of phosphatidylinositol bisphosphate (PIP2) in schizont-stage parasites, underlining its critical role in regulating phosphoinositide levels in the parasite. IMPORTANCE The clinical symptoms of malaria arise due to repeated rounds of replication of Plasmodium parasites within red blood cells (RBCs). Central to this is an intense period of membrane biogenesis. Generation of membranes not only requires de novo synthesis and acquisition but also the degradation of phospholipids, a function that is performed by phospholipases. In this study, we investigate the essentiality of the 19 putative phospholipase enzymes that the human malaria parasite Plasmodium falciparum expresses during its replication within RBCs. We not only show that a high level of functional redundancy exists among these enzymes but, at the same time, also identify an essential role for the phosphoinositide-specific phospholipase C in parasite development and cleavage of the phospholipid phosphatidylinositol bisphosphate.

Longitudinal gut microbiome analyses and blooms of pathogenic strains during lupus disease flares.

OBJECTIVE: Whereas genetic susceptibility for systemic lupus erythematosus (SLE) has been well explored, the triggers for clinical disease flares remain elusive. To investigate relationships between microbiota community resilience and disease activity, we performed the first longitudinal analyses of lupus gut-microbiota communities. METHODS: In an observational study, taxononomic analyses, including multivariate analysis of ß-diversity, assessed time-dependent alterations in faecal communities from patients and healthy controls. From gut blooms, strains were isolated, with genomes and associated glycans analysed. RESULTS: Multivariate analyses documented that, unlike healthy controls, significant temporal community-wide ecological microbiota instability was common in SLE patients, and transient intestinal growth spikes of several pathogenic species were documented. Expansions of only the anaerobic commensal, Ruminococcus (blautia) gnavus (RG) occurred at times of high-disease activity, and were detected in almost half of patients during lupus nephritis (LN) disease flares. Whole genome sequence analysis of RG strains isolated during these flares documented 34 genes postulated to aid adaptation and expansion within a host with an inflammatory condition. Yet, the most specific feature of strains found during lupus flares was the common expression of a novel type of cell membrane-associated lipoglycan. These lipoglycans share conserved structural features documented by mass spectroscopy, and highly immunogenic repetitive antigenic-determinants, recognised by high-level serum IgG2 antibodies, that spontaneously arose, concurrent with RG blooms and lupus flares. CONCLUSIONS: Our findings rationalise how blooms of the RG pathobiont may be common drivers of clinical flares of often remitting-relapsing lupus disease, and highlight the potential pathogenic properties of specific strains isolated from active LN patients.

A Single Residue within the MCR-1 Protein Confers Anticipatory Resilience.

The envelope stress response (ESR) of Gram-negative enteric bacteria senses fluctuations in nutrient availability and environmental changes to avert damage and promote survival. It has a protective role toward antimicrobials, but direct interactions between ESR components and antibiotic resistance genes have not been demonstrated. Here, we report interactions between a central regulator of ESR viz., the two-component signal transduction system CpxRA (conjugative pilus expression), and the recently described mobile colistin resistance protein (MCR-1). Purified MCR-1 is specifically cleaved within its highly conserved periplasmic bridge element, which links its N-terminal transmembrane domain with the C-terminal active-site periplasmic domain, by the CpxRA-regulated serine endoprotease DegP. Recombinant strains harboring cleavage site mutations in MCR-1 are either protease resistant or degradation susceptible, with widely differing consequences for colistin resistance. Transfer of the gene encoding a degradation-susceptible mutant to strains that lack either DegP or its regulator CpxRA restores expression and colistin resistance. MCR-1 production in Escherichia coli imposes growth restriction in strains lacking either DegP or CpxRA, effects that are reversed by transactive expression of DegP. Excipient allosteric activation of the DegP protease specifically inhibits growth of isolates carrying mcr-1 plasmids. As CpxRA directly senses acidification, growth of strains at moderately low pH dramatically increases both MCR-1-dependent phosphoethanolamine (PEA) modification of lipid A and colistin resistance levels. Strains expressing MCR-1 are also more resistant to antimicrobial peptides and bile acids. Thus, a single residue external to its active site induces ESR activity to confer resilience in MCR-1-expressing strains to commonly encountered environmental stimuli, such as changes in acidity and antimicrobial peptides. Targeted activation of the nonessential protease DegP can lead to the elimination of transferable colistin resistance in Gram-negative bacteria. IMPORTANCE The global presence of transferable mcr genes in a wide range of Gram-negative bacteria from clinical, veterinary, food, and aquaculture environments is disconcerting. Its success as a transmissible resistance factor remains enigmatic, because its expression imposes fitness costs and imparts only moderate levels of colistin resistance. Here, we show that MCR-1 triggers regulatory components of the envelope stress response, a system that senses fluctuations in nutrient availability and environmental changes, to promote bacterial survival in low pH environments. We identify a single residue within a highly conserved structural element of mcr-1 distal to its catalytic site that modulates resistance activity and triggers the ESR. Using mutational analysis, quantitative lipid A profiling and biochemical assays, we determined that growth in low pH environments dramatically increases colistin resistance levels and promotes resistance to bile acids and antimicrobial peptides. We exploited these findings to develop a targeted approach that eliminates mcr-1 and its plasmid carriers.

Resolving colistin resistance and heteroresistance in Enterobacter species.

Species within the Enterobacter cloacae complex (ECC) include globally important nosocomial pathogens. A three-year study of ECC in Germany identified Enterobacter xiangfangensis as the most common species (65.5%) detected, a result replicated by examining a global pool of 3246 isolates. Antibiotic resistance profiling revealed widespread resistance and heteroresistance to the antibiotic colistin and detected the mobile colistin resistance (mcr)-9 gene in 19.2% of all isolates. We show that resistance and heteroresistance properties depend on the chromosomal arnBCADTEF gene cassette whose products catalyze transfer of L-Ara4N to lipid A. Using comparative genomics, mutational analysis, and quantitative lipid A profiling we demonstrate that intrinsic lipid A modification levels are genospecies-dependent and governed by allelic variations in phoPQ and mgrB, that encode a two-component sensor-activator system and specific inhibitor peptide. By generating phoPQ chimeras and combining them with mgrB alleles, we show that interactions at the pH-sensing interface of the sensory histidine kinase phoQ dictate arnBCADTEF expression levels. To minimize therapeutic failures, we developed an assay that accurately detects colistin resistance levels for any ECC isolate.

Are n-3 PUFAs from Microalgae Incorporated into Membrane and Storage Lipids in Pig Muscle Tissues?-A Lipidomic Approach.

For the study of molecular mechanisms of to lipid transport and storage in relation to dietary effects, lipidomics has been rarely used in farm animal research. A feeding study with pigs (German Landrace sows) and supplementation of microalgae (Schizochytrium sp.) was conducted. The animals were allocated to the control group (n = 15) and the microalgae group (n = 16). Shotgun lipidomics was applied. This study enabled us to identify and quantify 336 lipid species from 15 different lipid classes in pig skeletal muscle tissues. The distribution of the lipid classes was significantly altered by microalgae supplementation, and ether lipids of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidic acid (PA) were significantly decreased. The total concentration of triacylglycerides (TAGs) was not affected. TAGs with high degree of unsaturation (TAG 56:7, TAG 56:6, TAG 54:6) were increased in the microalgae group, and major abundant species like TAG 52:2 and TAG 52:1 were not affected by the diet. Our results confirmed that dietary DHA and EPA are incorporated into storage and membrane lipids of pig muscles, which further led to systemic changes in the lipidome composition.

A Current Encyclopedia of Bioinformatics Tools, Data Formats and Resources for Mass Spectrometry Lipidomics.

Mass spectrometry is a widely used technology to identify and quantify biomolecules such as lipids, metabolites and proteins necessary for biomedical research. In this study, we catalogued freely available software tools, libraries, databases, repositories and resources that support lipidomics data analysis and determined the scope of currently used analytical technologies. Because of the tremendous importance of data interoperability, we assessed the support of standardized data formats in mass spectrometric (MS)-based lipidomics workflows. We included tools in our comparison that support targeted as well as untargeted analysis using direct infusion/shotgun (DI-MS), liquid chromatography-mass spectrometry, ion mobility or MS imaging approaches on MS1 and potentially higher MS levels. As a result, we determined that the Human Proteome Organization-Proteomics Standards Initiative standard data formats, mzML and mzTab-M, are already supported by a substantial number of recent software tools. We further discuss how mzTab-M can serve as a bridge between data acquisition and lipid bioinformatics tools for interpretation, capturing their output and transmitting rich annotated data for downstream processing. However, we identified several challenges of currently available tools and standards. Potential areas for improvement were: adaptation of common nomenclature and standardized reporting to enable high throughput lipidomics and improve its data handling. Finally, we suggest specific areas where tools and repositories need to improve to become FAIRer.

Tuberculostearic Acid-Containing Phosphatidylinositols as Markers of Bacterial Burden in Tuberculosis.

One-fourth of the global human population is estimated to be infected with strains of the Mycobacterium tuberculosis complex (MTBC), the causative agent of tuberculosis (TB). Using lipidomic approaches, we show that tuberculostearic acid (TSA)-containing phosphatidylinositols (PIs) are molecular markers for infection with clinically relevant MTBC strains and signify bacterial burden. For the most abundant lipid marker, detection limits of ∼102 colony forming units (CFUs) and ∼103 CFUs for bacterial and cell culture systems were determined, respectively. We developed a targeted lipid assay, which can be performed within a day including sample preparation─roughly 30-fold faster than in conventional methods based on bacterial culture. This indirect and culture-free detection approach allowed us to determine pathogen loads in infected murine macrophages, human neutrophils, and murine lung tissue. These marker lipids inferred from mycobacterial PIs were found in higher levels in peripheral blood mononuclear cells of TB patients compared to healthy individuals. Moreover, in a small cohort of drug-susceptible TB patients, elevated levels of these molecular markers were detected at the start of therapy and declined upon successful anti-TB treatment. Thus, the concentration of TSA-containing PIs can be used as a correlate for the mycobacterial burden in experimental models and in vitro systems and may prospectively also provide a clinically relevant tool to monitor TB severity.

Characterization of phospholipid-modified lung surfactant in vitro and in a neonatal ARDS model reveals anti-inflammatory potential and surfactant lipidome signatures.

A strong inflammatory immune response drives the lung pathology in neonatal acute respiratory distress syndrome (nARDS). Anti-inflammatory therapy is therefore a promising strategy for improved treatment of nARDS. We demonstrate a new function of the anionic phospholipids POPG, DOPG, and PIP2 as inhibitors of IL-1ß release by LPS and ATP-induced inflammasome activation in human monocyte-derived and lung macrophages. Curosurf® surfactant was enriched with POPG, DOPG, PIP2 and the head-group derivative IP3, biophysically characterized and applicability was evaluated in a piglet model of nARDS. The composition of pulmonary surfactant from piglets was determined by shotgun lipidomics screens. After 72 h of nARDS, levels of POPG, DOPG, and PIP2 were enhanced in the respective treatment groups. Otherwise, we did not observe changes of individual lipid species in any of the groups. Surfactant proteins were not affected, with the exception of the IP3 treated group. Our data show that POPG, DOPG, and PIP2 are potent inhibitors of inflammasome activation; their enrichment in a surfactant preparation did not induce any negative effects on lipid profile and reduced biophysical function in vitro was mainly observed for PIP2. These results encourage to rethink the current strategies of improving surfactant preparations by inclusion of anionic lipids as potent anti-inflammatory immune regulators.

Sub-Lineage Specific Phenolic Glycolipid Patterns in the Mycobacterium tuberculosis Complex Lineage 1.

"Ancestral" Mycobacterium tuberculosis complex (MTBC) strains of Lineage 1 (L1, East African Indian) are a prominent tuberculosis (TB) cause in countries around the Indian Ocean. However, the pathobiology of L1 strains is insufficiently characterized. Here, we used whole genome sequencing (WGS) of 312 L1 strains from 43 countries to perform a characterization of the global L1 population structure and correlate this to the analysis of the synthesis of phenolic glycolipids (PGL) - known MTBC polyketide-derived virulence factors. Our results reveal the presence of eight major L1 sub-lineages, whose members have specific mutation signatures in PGL biosynthesis genes, e.g., pks15/1 or glycosyltransferases Rv2962c and/or Rv2958c. Sub-lineage specific PGL production was studied by NMR-based lipid profiling and strains with a completely abolished phenolphthiocerol dimycoserosate biosynthesis showed in average a more prominent growth in human macrophages. In conclusion, our results show a diverse population structure of L1 strains that is associated with the presence of specific PGL types. This includes the occurrence of mycoside B in one sub-lineage, representing the first description of a PGL in an M. tuberculosis lineage other than L2. Such differences may be important for the evolution of L1 strains, e.g., allowing adaption to different human populations.

The human LL-37 peptide exerts antimicrobial activity against Legionella micdadei interacting with membrane phospholipids.

Legionella micdadei is responsible for community- or nosocomial-acquired pneumonia as well as the influenza-like illness Pontiac fever. The aim of this study was to investigate the ability of L. micdadei to utilize extracellular choline for phosphatidylcholine (PC) synthesis and its consequences for the phospholipid composition of its membrane system and the interaction with the human LL-37 peptide. Comparative analysis of the PC content using isotopic labeling revealed that in presence of exogenous choline 98% of the total PC was synthesized via the Pcs pathway while the remaining 2% were generated via the PE-methylation (PmtA) pathway. PC species were to a greater extent defined by the Pcs pathway in the outer membrane than in the inner membrane. While no major changes in the bacterial lipid content were observed using 31P NMR, indication for utilization of longer acyl chains and slight increase of PG in response to choline addition was observed by a top-down lipidomics screen. The LL-37 peptide inhibited L. micdadei growth in a dose-dependent manner. Bacteria cultured with exogenous choline were more sensitive to the LL-37 peptide when compared to the standard culture condition. Our biophysical investigations show that the peptide perturbs bacterial-derived phospholipid monolayers and this interaction is dependent on the molar portion of PC. This interaction is responsible for the observed changes in the anti-L. micdadei activity of the LL-37 peptide.

Nano-in-Microparticles for Aerosol Delivery of Antibiotic-Loaded, Fucose-Derivatized, and Macrophage-Targeted Liposomes to Combat Mycobacterial Infections: In Vitro Deposition, Pulmonary Barrier Interactions, and Targeted Delivery.

Nontuberculous mycobacterial infections rapidly emerge and demand potent medications to cope with resistance. In this context, targeted loco-regional delivery of aerosol medicines to the lungs is an advantage. However, sufficient antibiotic delivery requires engineered aerosols for optimized deposition. Here, the effect of bedaquiline-encapsulating fucosylated versus nonfucosylated liposomes on cellular uptake and delivery is investigated. Notably, this comparison includes critical parameters for pulmonary delivery, i.e., aerosol deposition and the noncellular barriers of pulmonary surfactant (PS) and mucus. Targeting increases liposomal uptake into THP-1 cells as well as peripheral blood monocyte- and lung-tissue derived macrophages. Aerosol deposition in the presence of PS, however, masks the effect of active targeting. PS alters antibiotic release that depends on the drug's hydrophobicity, while mucus reduces the mobility of nontargeted more than fucosylated liposomes. Dry-powder microparticles of spray-dried bedaquiline-loaded liposomes display a high fine particle fraction of >70%, as well as preserved liposomal integrity and targeting function. The antibiotic effect is maintained when deposited as powder aerosol on cultured Mycobacterium abscessus. When treating M. abscessus infected THP-1 cells, the fucosylated variant enabled enhanced bacterial killing, thus opening up a clear perspective for the improved treatment of nontuberculous mycobacterial infections.

Remodeling of Lipid A in Pseudomonas syringae pv. phaseolicola In Vitro.

Pseudomonas species infect a variety of organisms, including mammals and plants. Mammalian pathogens of the Pseudomonas family modify their lipid A during host entry to evade immune responses and to create an effective barrier against different environments, for example by removal of primary acyl chains, addition of phosphoethanolamine (P-EtN) to primary phosphates, and hydroxylation of secondary acyl chains. For Pseudomonas syringae pv. phaseolicola (Pph) 1448A, an economically important pathogen of beans, we observed similar lipid A modifications by mass spectrometric analysis. Therefore, we investigated predicted proteomes of various plant-associated Pseudomonas spp. for putative lipid A-modifying proteins using the well-studied mammalian pathogen Pseudomonas aeruginosa as a reference. We generated isogenic mutant strains of candidate genes and analyzed their lipid A. We show that the function of PagL, LpxO, and EptA is generally conserved in Pph 1448A. PagL-mediated de-acylation occurs at the distal glucosamine, whereas LpxO hydroxylates the secondary acyl chain on the distal glucosamine. The addition of P-EtN catalyzed by EptA occurs at both phosphates of lipid A. Our study characterizes lipid A modifications in vitro and provides a useful set of mutant strains relevant for further functional studies on lipid A modifications in Pph 1448A.

Interleukin-13-Overexpressing Mice Represent an Advanced Preclinical Model for Detecting the Distribution of Antimycobacterial Drugs within Centrally Necrotizing Granulomas.

The Mycobacterium tuberculosis-harboring granuloma with a necrotic center surrounded by a fibrous capsule is the hallmark of tuberculosis (TB). For a successful treatment, antibiotics need to penetrate these complex structures to reach their bacterial targets. Hence, animal models reflecting the pulmonary pathology of TB patients are of particular importance to improve the preclinical validation of novel drug candidates. M. tuberculosis-infected interleukin-13-overexpressing (IL-13tg) mice develop a TB pathology very similar to patients and, in contrast to other mouse models, also share pathogenetic mechanisms. Accordingly, IL-13tg animals represent an ideal model for analyzing the penetration of novel anti-TB drugs into various compartments of necrotic granulomas by matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MS imaging). In the present study, we evaluated the suitability of BALB/c IL-13tg mice for determining the antibiotic distribution within necrotizing lesions. To this end, we established a workflow based on the inactivation of M. tuberculosis by gamma irradiation while preserving lung tissue integrity and drug distribution, which is essential for correlating drug penetration with lesion pathology. MALDI-MS imaging analysis of clofazimine, pyrazinamide, and rifampicin revealed a drug-specific distribution within different lesion types, including cellular granulomas, developing in BALB/c wild-type mice, and necrotic granulomas in BALB/c IL-13tg animals, emphasizing the necessity of preclinical models reflecting human pathology. Most importantly, our study demonstrates that BALB/c IL-13tg mice recapitulate the penetration of antibiotics into human lesions. Therefore, our workflow in combination with the IL-13tg mouse model provides an improved and accelerated evaluation of novel anti-TB drugs and new regimens in the preclinical stage.

WNT6/ACC2-induced storage of triacylglycerols in macrophages is exploited by Mycobacterium tuberculosis.

In view of emerging drug-resistant tuberculosis (TB), host-directed adjunct therapies are urgently needed to improve treatment outcomes with currently available anti-TB therapies. One approach is to interfere with the formation of lipid-laden "foamy" macrophages in the host, as they provide a nutrient-rich host cell environment for Mycobacterium tuberculosis (Mtb). Here, we provide evidence that Wnt family member 6 (WNT6), a ligand of the evolutionarily conserved Wingless/Integrase 1 (WNT) signaling pathway, promotes foam cell formation by regulating key lipid metabolic genes including acetyl-CoA carboxylase 2 (ACC2) during pulmonary TB. Using genetic and pharmacological approaches, we demonstrated that lack of functional WNT6 or ACC2 significantly reduced intracellular triacylglycerol (TAG) levels and Mtb survival in macrophages. Moreover, treatment of Mtb-infected mice with a combination of a pharmacological ACC2 inhibitor and the anti-TB drug isoniazid (INH) reduced lung TAG and cytokine levels, as well as lung weights, compared with treatment with INH alone. This combination also reduced Mtb bacterial numbers and the size of mononuclear cell infiltrates in livers of infected mice. In summary, our findings demonstrate that Mtb exploits WNT6/ACC2-induced storage of TAGs in macrophages to facilitate its intracellular survival, a finding that opens new perspectives for host-directed adjunctive treatment of pulmonary TB.