Biology subjects

Vidal, A. G.

Publications and source records attributed to Vidal, A. G..

6 recordsLinked to original sources

bioRxiv2026

Lactate Oxidase (LctO) Acts as a Metabolic Checkpoint Restraining Streptococcus pneumoniae Invasion of Respiratory Epithelial Barriers

Streptococcus pneumoniae rapidly translocates across polarized human bronchial epithelial barriers, with viable bacteria recovered from the basolateral compartment within 1 h post-infection. Disruption of the pyruvate node through combined deletion of pyruvate oxidase (spxB) and lactate oxidase (lctO) markedly enhanced transmigration of S. pneumoniae across polarized Calu-3 monolayers without causing early cytotoxicity or loss of monolayer integrity. This hyper-invasive phenotype was conserved in the TIGR4 and EF3030 background and under air-liquid interface conditions. Importantly, single {Delta}lctO mutants exhibited significantly greater translocation than {Delta}spxB mutants or wild-type strains across bronchial (Calu-3), alveolar (A549), and pharyngeal (Detroit 562) epithelial models. Enhanced translocation correlated with increased bacterial adherence but was independent of capsule expression, extracellular H2O2 production, pneumolysin, or tight junction disruption, as evidenced by stable transepithelial electrical resistance (TEER), lack of caspase-3/7 activation, and minimal IL-18 release at early time points. High-resolution confocal microscopy revealed intracellular {Delta}lctO pneumococci localized within N-acetylglucosamine/sialic acid (GN/SA)-containing compartments as early as 1 h post-infection. In murine macrophages, {Delta}lctO mutants were phagocytosed at rates similar to wild-type bacteria but induced greater pneumolysin-dependent cytotoxicity at 24 h. These findings demonstrate that LctO functions as a metabolic checkpoint that restrains pneumococcal invasion of respiratory epithelia, revealing a previously unrecognized role for lactate oxidase in controlling the transition from colonization to invasive disease. ImportanceThis study identifies lactate oxidase (LctO) as a critical metabolic checkpoint that restrains Streptococcus pneumoniae invasion of respiratory epithelial barriers. By linking pyruvate node metabolism to the control of transmigration, these findings reveal a novel mechanism by which central carbon metabolism regulates pneumococcal virulence and the transition from colonization to invasive disease.

microbiology↗

bioRxiv2026

A pan-serotype human monoclonal antibody protects against pneumococcal infection by targeting multiple choline binding domain proteins

Streptococcus pneumoniae remains a global health threat, particularly to young children, the elderly, and immunocompromised individuals. Pneumococcal vaccines targeting the bacterial capsule polysaccharide do not protect against all 100+ pneumococcal serotypes, contributing to non-vaccine serotype infections and antibiotic resistance. To address these limitations, we isolated human monoclonal antibodies (mAbs) targeting pneumococcal surface proteins and identified a first-in-class mAb, derived from a patient with prior pneumococcal infection, namely mAb 5995-40. mAb 5995-40 bound multiple pneumococcal proteins, including PcpA and PspA, through a conserved choline-binding domain shared across serotypes. Functionally, mAb 5995-40 provided complete protection in lethal pneumococcal challenge models and improved survival in influenza A, influenza B, and respiratory syncytial virus-associated bacterial coinfection models. Mechanistic studies showed enhanced opsonophagocytic killing, reduced bacterial dissemination, and blocked epithelial translocation. Cryo-electron microscopy identified a repeating motif within the choline-binding domain targeted by mAb 5995-40, highlighting its potential as a broadly protective pneumococcal therapeutic.

immunology↗

bioRxiv2025

Pneumococcal H2O2 Reshapes Mitochondrial Function and Reprograms Host Cell Metabolism

Streptococcus pneumoniae (Spn), a primary cause of pneumonia, induces acute lung parenchymal damage through a unique metabolic pathway generating hydrogen peroxide (H2O2) as a byproduct. This study demonstrates that Spn-derived H2O2, primarily produced by pyruvate oxidase (SpxB), inhibits key tricarboxylic acid (TCA) cycle enzymes (aconitase, glutamate dehydrogenase, and -ketoglutarate dehydrogenase) in lung epithelial cells, leading to citrate accumulation and diminished NADH production for oxidative phosphorylation. RNA sequencing reveals SpxB-dependent upregulation of glycolytic genes (HIF1A, IER3, HK2, PFKP), restricting pyruvate entry into the TCA cycle and increasing glucose consumption and lactate/acetate production, indicative of a Warburg-like metabolic shift that may enhance bacterial survival. Notably, mitochondrial membrane potential remains largely preserved, with minimal apoptosis despite Spn-induced stress. These findings uncover a novel mechanism of Spn-driven host metabolic reprogramming, highlighting potential therapeutic targets for pneumococcal diseases. ImportanceStreptococcus pneumoniae remains a leading cause of community-acquired pneumonia worldwide, yet the mechanisms by which it manipulates host metabolism to promote its survival and pathogenesis are not fully understood. This study reveals a novel metabolic strategy whereby pneumococcus-derived hydrogen peroxide, generated by pyruvate oxidase (SpxB), disrupts the host TCA cycle and drives a Warburg-like metabolic shift in lung epithelial cells. By inhibiting key TCA cycle enzymes and rewiring glycolytic gene expression, S. pneumoniae effectively reprograms host cell metabolism to favor its persistence, while minimizing host cell apoptosis and maintaining mitochondrial function. These insights expand our understanding of host-pathogen metabolic interactions and identify potential metabolic vulnerabilities that could be targeted to mitigate tissue damage and improve treatment outcomes in pneumococcal pneumonia.

microbiology↗

bioRxiv2023

Ultrastructural, metabolic and genetic determinants of the acquisition of macrolide resistance by Streptococcus pneumoniae

AimStreptococcus pneumoniae (Spn) acquires genes for macrolide resistance, MEGA or ermB, in the human host. These genes are carried either in the chromosome, or on integrative conjugative elements (ICEs). Here, we investigated molecular determinants of the acquisition of macrolide resistance. Methods and ResultsWhole genome analysis was conducted for 128 macrolide-resistant pneumococcal isolates to identify the presence of MEGA (44.5%, 57/128) or ermB (100%), and recombination events in Tn916-related elements or in the locus comCDE encoding competence genes. Confocal and electron microscopy studies demonstrated that, during the acquisition of macrolide resistance, pneumococcal strains formed clusters of varying size, with the largest aggregates having a median size of [~]1600 {micro}m2. Remarkably, these pneumococcal aggregates comprise both encapsulated and nonencapsulated pneumococci, exhibited physical interaction, and spanned extracellular and intracellular compartments. We assessed the recombination frequency (rF) for the acquisition of macrolide resistance by a recipient D39 strain, from pneumococcal strains carrying MEGA ([~]5.4 kb) in the chromone, or in large ICEs (>23 kb). Notably, the rF for the acquisition of MEGA, whether in the chromosome or carried on an ICE was similar. However, the rF adjusted to the acquisition of the full-length ICE ([~]52 kb), compared to that of the capsule locus ([~]23 kb) that is acquired by transformation, was three orders of magnitude higher. Finally, metabolomics studies revealed a link between the acquisition of ICE and the metabolic pathways involving nicotinic acid and sucrose. ConclusionsExtracellular and intracellular pneumococcal clusters facilitate the acquisition of full-length ICE at a rF higher than that of typical transformation events, involving distinct metabolic changes that present potential targets for interventions.

microbiology↗

bioRxiv2023

Oxidation of hemoproteins by Streptococcus pneumoniae collapses the cell cytoskeleton and disrupts mitochondrial respiration leading to cytotoxicity of human lung cells

Streptococcus pneumoniae (Spn) causes pneumonia that kills millions through acute toxicity and invasion of the lung parenchyma. During aerobic respiration, Spn releases hydrogen peroxide (Spn-H2O2), as a by-product of enzymes SpxB and LctO, and causes cell death with signs of both apoptosis and pyroptosis by oxidizing unknown cell targets. Hemoproteins are molecules essential for life and prone to oxidation by H2O2. We recently demonstrated that during infection-mimicking conditions, Spn-H2O2 oxidizes the hemoprotein hemoglobin (Hb), releasing toxic heme. In this study, we investigated details of the molecular mechanism(s) by which the oxidation of hemoproteins by Spn-H2O2 causes human lung cell death. Spn strains, but not H2O2-deficient Spn{Delta}spxB{Delta}lctO strains caused time-dependent cell cytotoxicity characterized by the rearrangement of the actin, the loss of the microtubule cytoskeleton and nuclear contraction. Disruption of the cell cytoskeleton correlated with the presence of invasive pneumococci and an increase of intracellular reactive oxygen species. In cell culture, the oxidation of Hb or cytochrome c (Cytc) caused DNA degradation and mitochondrial dysfunction from inhibition of complex I-driven respiration, which was cytotoxic to human alveolar cells. Oxidation of hemoproteins resulted in the creation of a radical, which was identified as a protein derived side chain tyrosyl radical by using electron paramagnetic resonance (EPR). Thus, we demonstrate that Spn invades lung cells, releasing H2O2 that oxidizes hemoproteins, including Cytc, catalyzing the formation of a tyrosyl side chain radical on Hb and causing mitochondrial disruption, that ultimately leads to the collapse of the cell cytoskeleton.

microbiology↗

bioRxiv2021

Prophylactic inhibition of colonization by Streptococcus pneumoniae with the secondary bile acid metabolite deoxycholic acid

Streptococcus pneumoniae (Spn) colonizes the nasopharynx of children and the elderly but also kills millions worldwide yearly. The secondary bile acid metabolite, deoxycholic acid (DoC), affects the viability of human pathogens but also plays multiple roles in host physiology. We assessed in vitro the antimicrobial activity of DoC and investigated its potential to eradicate Spn colonization using an ex vivo model of human nasopharyngeal colonization and an in vivo mouse model of colonization. At a physiological concentration DoC (0.5 mg/ml; 1.27 mM) killed all tested Spn strains (N=48) two h post-inoculation. The ex-vivo model of nasopharyngeal colonization showed that DoC eradicated colonization by Spn strains as soon as 10 min post-exposure. The mechanism of action did not involve activation of autolysis since the autolysis-defective double mutants {Delta}lytA{Delta}lytC and{Delta} spxB{Delta}lctO were as susceptible to DoC as was the wild-type (WT). Oral streptococcal species (N=20), however, were not susceptible to DoC (0.5 mg/ml). Unlike trimethoprim, whose spontaneous resistance frequency (srF) for TIGR4 or EF3030 was [≥]1x10-9, no spontaneous resistance was observed with DoC (srF[≥]1x10-12). Finally, the efficacy of DoC to eradicate Spn colonization was assessed in vivo using a topical route via intranasal (i.n.) administration and as a prophylactic treatment. Mice challenged with Spn EF3030 carried a median of 4.05x105 cfu/ml four days post-inoculation compared to 6.67x104 cfu/ml for mice treated with DoC. Mice in the prophylactic group had a [~]99% reduction of the pneumococcal density (median, 2.61 x103 cfu/ml). Thus, DoC, an endogenous human bile salt, has therapeutic potential against Spn.

microbiology↗