Biology subjects

David, L.

Publications and source records attributed to David, L..

At least 19 recordsLinked to original sources

bioRxiv2026

Recapitulating the notochord to nucleus pulposus transition from human mesendoderm progenitors in 3D

Intervertebral disc (IVD) degeneration, a major cause of low back pain, is driven by the disruption of nucleus pulposus (NP) tissue homeostasis. The NP arises from embryonic notochord cells that differentiate and adapt to the hypoxic, matrix-rich environment of the developing disc. Recapitulating the developmental trajectory from notochordal progenitors to NP-like cells in vitro remains challenging, as most protocols bypass early specification steps and fail to capture this understudied lineage. Here, using a three-dimensional (3D) system, we reconstruct this developmental path from human pluripotent stem cells into mesendoderm progenitor cells (MEPCs), notochord-like cells (NLCs), and NP-like cells, guided by single-cell transcriptomics and spatial profiling. We identify MEPCs as a distinct progenitor population with a unique gene signature and conserved regulatory networks (GSC, KLF8, LHX1, MIXL1) shared with early organizer and mesendodermal cells. Self-organization in MEPCs-derived aggregates induced spontaneous expression of NOTO, a key transcription factor orchestrating notochordal identity, and enhanced commitment to notochordal fate. Long-term culture in a medium containing TGF-{beta}1 and GDF-5 promoted maturation into SOX9+, KRT8+, CD109+ NP-like cells exhibiting robust matrix deposition and prominent LAMP1-positive intracytoplasmic vacuoles. This scalable, physiologically relevant model enables fundamental studies of human disc biology and early degenerative processes, while advancing regenerative strategies.

cell biology↗

bioRxiv2026

Hydrogel confinement enables subcutaneous delivery of vesicant antibody-drug conjugates

Antibody-drug conjugates (ADCs) deliver cytotoxic payloads to tumors with antibody selectivity, yet all approved ADCs are administered by intravenous (IV) infusion despite a strong patient and clinical preference for subcutaneous (SC) delivery. SC administration would reduce treatment burden, but many ADC payloads are vesicants that cause tissue necrosis upon local release, a liability amplified, not mitigated, by the dispersion-enhancing excipients used for SC antibody formulations. We developed an injectable diacetyl-L-tartaric anhydride-functionalized chitosan hydrogel (TACT) that addresses this conflict by confining ADCs within a protective SC depot. TACT is compatible with clinically approved ADC formulations without drug-product modification and provides drug-to-antibody ratio (DAR)-dependent release kinetics that support a quantitative relationship with in vivo absorption timing. In direct comparison, recombinant human hyaluronidase (rHuPH20) co-formulated with vesicant ADCs caused severe tissue necrosis, whereas TACT prevented macroscopic injury while preserving antitumor efficacy comparable to intravenous dosing. TUNEL staining of injection sites showed that TACT attenuated peri-depot apoptotic injury 3-fold relative to T-DM1 alone and 2-fold relative to rHuPH20 co-formulation. In non-human primates, SC TACT achieved 78% relative bioavailability for total trastuzumab, reduced peak circulating T-DM1 catabolite (free DM1) exposure 7.6-fold compared to IV administration and produced only transient, self-resolving cutaneous reactions. These results identify depot-mediated confinement as a viable alternative to excipient-mediated dispersion for SC delivery of vesicant ADCs, demonstrated here for trastuzumab-based conjugates across two approved ADC drug products (T-DM1 and T-DXd, with non-cleavable MCC and cleavable peptide linkers), with supporting validation in a custom cleavable monomethyl auristatin E (MMAE) series. Additional validation with enfortumab vedotin (EV), a Nectin-4-targeting MMAE ADC, supported the applicability of this strategy beyond trastuzumab-based conjugates.

cancer biology↗

bioRxiv2026

Characterization of programmed cell death pathways activated in Mycobacterium tuberculosis-infected human macrophages

Mycobacterium tuberculosis (Mtb) primarily infects human lung macrophages, which serve as its major replication niche. Mtb can manipulate host macrophage cell death pathways to its advantage by inhibiting apoptosis and inducing necrotic cell death. However, the specific necrotic cell death pathway activated in human macrophages after Mtb infection remains unclear. Here, we used the THP-1 cell line and primary human monocyte-derived macrophage (hMDM) to analyze multiple programmed cell death pathways during days 1-3 after Mtb infection. Confocal microscopic analysis demonstrates that Mtb-infected THP-1 cells or hMDMs rarely exhibited apoptosis. Immunoblotting shows that Mtb induces significant CASP3 and GSDME activation in THP-1 cells, but not in hMDMs. We show that Mtb, in THP-1 cells but not hMDM, induces a significant increase in GSDMD cleavage, a hallmark of pyroptosis. MLKL phosphorylation was not observed in THP-1 cells or hMDMs during Mtb infections, indicating an absence of necroptosis. No changes in ferroptosis markers such as GPX4 expression or lipid peroxidation levels were detected. Time-lapse live-cell imaging revealed no lysosomal membrane permeabilization prior to plasma membrane rupture (PMR). However, we observed DNA release from Mtb-infected THP-1 cells and hMDMs after PMR. The DNA released from THP-1 cells exhibits low levels of myeloperoxidase and histone H3 citrullination. High-resolution confocal imaging shows that Mtb is associated with the released DNA. We demonstrate that pyroptosis induction in THP-1 cells is dispensable for the DNA release and cell death induction. In conclusion, our results reveal that Mtb-triggered cell death in hMDMs bypasses canonical cell death pathways like apoptosis, pyroptosis, necroptosis, and ferroptosis. Instead, cell death in both THP-1 cells and hMDMs correlates with DNA release, potentially through a pathway similar to NETosis in neutrophils.

cell biology↗

bioRxiv2025

SMARCA2/4-Dependent Chromatin Remodelling Establishes Gene Regulatory Programs in Early Human Embryos and Blastoids

Establishment of cell lineages during development is regulated by transcription factors binding at cis-regulatory elements to activate gene regulatory programs. Many transcription factors require chromatin remodellers for accessibility at their target sites. Although these key principles of gene regulation in mammalian development have emerged, the contribution of chromatin remodellers to early human embryogenesis remains unknown. Here, we show that the SWI/SNF ATPases SMARCA2 and SMARCA4 are required for establishing the epiblast and trophectoderm fates during human pre-implantation development and facilitate accessibility at regulatory elements. We find that degradation of SWI/SNF ATPases disrupts epiblast formation in blastoids and enhances trophectoderm specification, while also showing transcriptional and chromatin misregulation in TE-like cells. In human embryos, SWI/SNF perturbation impaired blastocyst formation and the establishment of the inner cell mass. Single-nucleus chromatin accessibility and transcriptome profiling in blastoids reveals that the SWI/SNF complex safeguards the naive epiblast and trophectoderm programs and facilitates enhancer and transcription factor motif accessibility. These findings identify SWI/SNF chromatin remodellers as critical regulators of embryonic lineage specification during human pre-implantation development.

developmental biology↗

bioRxiv2025

Decoding the pituitary gonadotrope regulatory architecture governing the preovulatory surge in vivo

Despite the global burden of female infertility, the molecular control of ovulation remains unclear. Pituitary gonadotropes tune hormonal output to regulate ovarian function, yet the molecular mechanisms that control their function during the sexual cycle remain poorly understood. Using state-of-the-art single-nucleus multiomic profiling of the female rat pituitary across the estrus cycle, we uncover unexpected cyclic epigenetic and transcriptional remodelling in gonadotrope cells, particularly during the preovulatory surge. This plasticity is driven by a biphasic gene regulatory network switch that orchestrates large-scale reorganisation of the secretory machinery. We further demonstrate that, contrary to the prevailing model, the LH surge does not depend on Lhb transcriptional upregulation. Instead, the FSH surge arises from dynamic Fshb transcription mediated by a complex combination of newly identified enhancers and key transcription factors. Overall, our findings define a new molecular framework for gonadotropin surge regulation and provide a foundational resource for understanding female reproductive disorders.

physiology↗

bioRxiv2025

A deep learning approach for rational affinity maturation of anti-VEGF nanobodies

Nanobodies offer several advantages over conventional antibodies due to their lower immunogenicity, enhanced stability, and superior tissue penetration, making them promising candidates for cancer therapy. In this study, we employ deep learning algorithms to design anti-VEGF nanobodies via affinity maturation. Our approach integrates structure-guided mutational modeling and systematic measurement of binding affinity and stability for rational optimization of Complementarity Determining Regions. In addition, we developed a sequence-based melting temperature predictor for nanobodies, ensuring stability of the designed mutants. Our method achieves energy reductions up to -4.92 kcal/mol. Our melting temperature predictor demonstrated a Pearson correlation coefficient of 0.772. These findings emphasize the potential of computational approaches for nanobody affinity maturation and stability prediction, paving the way for more effective therapeutic designs.

bioinformatics↗

bioRxiv2025

Patient-derived Brain Organoids Reveal Divergent Neuronal Activity Across Subpopulations of Autism Spectrum Disorder

Patient-derived brain organoids have emerged as a powerful model for investigating the mechanisms underlying neurological and psychiatric disorders. They provide novel insights into autism spectrum disorder (ASD), a heterogeneous neurodevelopmental condition whose underlying mechanisms remain poorly understood. Recent advancements in generating electrophysiological functional 3D brain organoids enable the study of molecular and network-level neuronal activity. Here, we aimed to characterize the neurophysiological underpinnings of ASD by comparing electrophysiological properties of brain organoids derived from eleven individuals diagnosed with autism spectrum disorder -10 with monogenic syndromic ASD across five genetic subtypes, and 1 with idiopathic ASD -to organoids derived from 4 neurotypical control individuals. We identified distinct differences in baseline activity (resting state) and evoked responses (synaptic plasticity and network dynamics) across ASD subgroups. To comprehensively assess these differences, we applied dimensionality reduction and machine learning (principal component analysis, PCA) to integrate multiple electrophysiological features into a unified framework. Our findings reveal subtype-specific neurophysiological alterations in ASD brain organoids, offering mechanistic insights into ASD heterogeneity and potential applications for early diagnostics, drug screening, and therapeutic development.

neuroscience↗

bioRxiv2025

Identification of novel candidate regulators of human naive pluripotency by means of a genetic switch utilizing the chimeric receptor G-CSFR:GP130

This study investigated the molecular mechanisms underpinning the transition from primed to naive pluripotency in human pluripotent stem cells (hPSCs) by utilizing a dual genetic switch combining hormone-dependent STAT3-ERT2 and a chimeric GCSFR:gp130 receptor. Upon activation with tamoxifen and G-CSF, hPSCs demonstrated upregulation of naive markers, and sustained self-renewal independent of LIF. Transcriptomic analyses revealed distinct clusters of early response genes, with a subset of 26 genes significantly enriched in the human epiblast. Knockdown experiments further delineated essential regulators that modulate differentiation and proliferation during naive reprogramming including interferon gamma inducible protein (IFI)16 and interferon-induced transmembrane (IFITM) proteins. Moreover, functional assays using mutant chimeric GCSFR:gp130 receptor highlighted the critical role of JAK kinase recruitment in orchestrating epigenomic remodeling and early gene activation. This approach establishes a robust platform to dissect the signaling dynamics governing the accession of hPSCs to naive pluripotency.

cell biology↗

bioRxiv2025

Outer Membrane Vesicles from bacteria expressing the HlyF/CprA family of enzymes are more efficient at delivering their cargo into host cells.

HlyF, a cytoplasmic short-chain dehydrogenase/reductase enzyme encoded by a virulence plasmid in Escherichia coli, and its ortholog CprA, encoded notably by the chromosome of Pseudomonas aeruginosa, drive the production of specific outer membrane vesicles (OMVs). These OMVs have been shown to have the unique abilities to disrupt autophagic flux and exacerbate inflammatory response in host cells. In this study, we demonstrate that these OMVs have improved cargo delivery efficiency through a clathrin-independent internalization pathway into host cells, by directly fusing with the plasma membrane of the host cell. These results reveal an uncovered role of the HlyF/CprA enzyme family in enhancing OMV function through a unique internalization pathway and unveil a novel mechanism driving OMV-mediated virulence.

microbiology↗

bioRxiv2025

HUMESS: Integrating Quantitative Transcriptomic Analysis and Metabolic Modeling to Unveil Condition-Specific Gene Signatures

Transcriptomic analysis is a key tool for exploring gene expression, but the complexity of biological systems often limits its insights. In particular, the lack of intermodal or multi-layered analysis hinders the ability to fully capture key cellular functions such as metabolism from transcriptomic data alone. Here, we introduce a novel approach that integrates transcriptomic data with metabolic network modeling to address this. Unlike traditional methods, HUMESS prioritizes genes based on their metabolic significance, offering a deeper understanding of condition-specific gene expression. Our computational pipeline, supported by a user-friendly Rshiny application, enhances gene expression analysis by uncovering metabolic phenotypic signatures.

systems biology↗

bioRxiv2024

NINJ1 is activated by calcium-driven plasma membrane lipid scrambling during lytic cell death

NINJ1 is the terminal executioner of cellular rupture in multiple lytic cell death pathways through its clustering in the plasma membrane. Its activation trigger, however, remains unknown. We found that NINJ1-mediated plasma membrane rupture depends on calcium influx into the cell, which suffices to induce NINJ1-mediated rupture. Using genetic and pharmacologic approaches in macrophages, we show calcium drives membrane rupture through phospholipid scrambling by the calcium-activated scramblase TMEM16F. We next tested whether this calcium-activated NINJ1 mechanism is the elusive pathway by which extracellular ATP stimulates cellular rupture. We show that ATP-stimulation of P2X7R induces NINJ1-mediated cell lysis via calcium influx and TMEM16F lipid scrambling, independently of inflammasomes, pannexins and gasdermin D. Our work reveals the mechanism of NINJ1 activation and solves the long-standing mystery of ATP-induced cytolysis. SummaryElevated cytosolic calcium drives NINJ1-mediated cellular rupture during lytic cell death through plasma membrane lipid scrambling.

cell biology↗

bioRxiv2024

Fatiguing Exercise Reduces Cellular Passive Young's Modulus in Human Vastus Lateralis Muscle

Previous studies demonstrated that acute, exercise-induced fatigue transiently reduces whole-muscle stiffness. Because reduced muscle stiffness at fatigue may contribute to increased injury risk and impaired contractile performance, the present study seeks to elucidate potential intracellular mechanisms underlying these reductions. To that end, cellular passive Youngs Modulus was measured in single, permeabilized muscle fibers from healthy, recreationally-active males and females. Eight volunteers (4 male, 4 female) completed unilateral, repeated maximal voluntary knee extensions until fatigue, after which percutaneous needle biopsies were performed on the fatigued (F) and non-fatigued (NF) Vastus Lateralis muscles. Muscle samples were processed for mechanical assessment and separately for imaging and phosphoproteomics. Single fibers were passively (pCa 8.0), incrementally stretched to 156% of the initial sarcomere length to assess Youngs Modulus, calculated as the slope of the resulting stress-strain curve at short (strain = 1.00-1.24 %Lo) and long (strain = 1.32-1.56 %Lo) fiber lengths. Titin phosphorylation was assessed by liquid chromatography followed by high-resolution mass spectrometry (LC-MS). Passive modulus was significantly reduced by fatigue at short and long lengths in male, but not female, participants. Fatigue increased phosphorylation of four serine residues located within the elastic region of titin and reduced phosphorylation at one serine residue but did not impact active tension nor sarcomere ultrastructure. Collectively, these results suggest muscle fatigue reduces cellular passive modulus in young males, but not females, concurrent with altered titin phosphorylation. These results provide mechanistic insight contributing to the understanding of sex-based differences in soft tissue injury and falls risk. Key Points SummaryO_LIPrevious studies have shown that skeletal muscle stiffness is reduced following a single bout of fatiguing exercise. C_LIO_LILower muscle stiffness at fatigue may increase risk for soft-tissue injury, however, the underlying mechanisms of this change are unclear. C_LIO_LIOur findings show that fatiguing exercise reduces passive Youngs modulus in skeletal muscle cells from males but not females, suggesting that intracellular proteins contribute to reduced muscle stiffness with fatigue in a sex-dependent manner. C_LIO_LIThe phosphorylation status of the intracellular protein titin is modified by fatiguing exercise in a way that may contribute to altered muscle stiffness after fatiguing exercise. C_LIO_LIThese results provide important mechanistic insight that may help explain why biological sex impacts risk for soft tissue injury in with repeated or high intensity mechanical loading in athletes and falls risk in older adults. C_LI New and NoteworthyMuscle fatigue has previously been shown to reduce musculotendinous stiffness, but the underlying mechanisms remain unclear. Our study presents novel evidence of fatigue-induced reductions in passive cellular Youngs Modulus in skeletal muscle from males, but not females, in conjunction with fatigue-induced alterations in titin phosphorylation. Collectively, these results suggest that intracellular mechanisms including titin phosphorylation may contribute to altered skeletal muscle stiffness following fatiguing exercise, and that this response is mediated by biological sex.

biophysics↗

bioRxiv2024

SMAC mimetics overcome apoptotic resistance in ovarian cancer through MSLN-TNF alpha axis

Resistance to chemotherapy and PARPi inhibitors remains a critical challenge in the treatment of epithelial ovarian cancer, mainly due to disabled apoptotic responses in tumor cells. Given mesothelins pivotal role in ovarian cancer and its restricted expression in healthy tissues, we conducted a drug-screening discovery analysis across a range of genetically modified cancer cells to unveil mesothelins therapeutic impact. We observed enhanced cell death in cancer cells with low mesothelin expression when exposed to a second mitochondria-derived activator of caspases (SMAC) mimetics, and demonstrated a compelling synergy when combined with chemotherapy in ex vivo patient-derived cultures and zebrafish tumor xenografts. Mechanistically, the addition of the SMAC mimetics drug birinapant to either carboplatin or paclitaxel triggered the activation of the Caspase 8-dependent apoptotic program facilitated by TNFLJ signaling. Multimodal analysis of neoadjuvant-treated patient samples further revealed an association between tumor-associated macrophages and the activation of TNFLJ-related pathways. Our proposed bimodal treatment shows promise in enhancing the clinical management of patients by harnessing the potential of SMAC mimetics alongside conventional chemotherapy.

cancer biology↗

bioRxiv2023

Combinatory differentiation of human induced pluripotent stem cells generates thymic epithelium that supports thymic crosstalk and directs dendritic- and CD4/CD8 T-cell full development

The thymus is a primary lymphoid organ playing a crucial role in immune tolerance, by educating thymocytes through a selection mediated by thymic epithelial cells (TEC). Recent advances in gene editing and immunotherapies have made in vitro generation of iPSC-derived T cells a crucial issue with promising therapeutic applications. Current state-of-the-art approaches often fail to accurately replicate the thymic niche, resulting in impaired T cell generation, particularly for CD4+ T cells. Here we address the production of functional mature iPSc-derived TECs that are able to support in vitro T cell generation. We designed a protocol allowing iPSc differentiation into thymic epithelial progenitors (TEP) through an unbiased multifactorial method based on optimal experimental design. Modulation of signaling pathways known to regulate embryonic thymus organogenesis resulted in the obtention of TEPs expressing typical thymic markers. We achieved TEP maturation into medullary and cortical TECs by setting up a hydrogel-based 3D culture supplemented in RANK ligand, resulting in the formation of thymic organoids. To assess the functionality of the generated TECs, primary hematopoietic progenitors were co-cultured in the organoids and showed significantly improved maturation in single-positive CD4 or CD8 T cells. Remarkably, our thymic organoid model shows multilineage differentiation potential, with generation of distinct dendritic cells populations in addition to the T lineages. Thus, generation of functional TECs and thymic organoids offers a practical platform for the study of thymic cellular interactions and paves the road to future cellular therapies by producing mature thymic immune cells of clinical interest.

immunology↗

bioRxiv2023

Single cell analysis reveals CD45RClow/-TNFR2+CD29lowCD8+ Tregs with superior activity

Although described in the 70s, CD8+ regulatory T cells (Tregs) remain incompletely understood and to date, although several markers are used to define them, they remain poorly defined. The identification of reliable and consistent markers, as it was done for CD4+ Tregs, remains an urgent task and a challenge to advance our understanding. Herein, we analyzed total CD8+ T cells using single cell CITEseq and VDJ T cell receptor sequencing utilizing markers used previously to identify Tregs, in particular CD45RC described by our team and others to divide pro-inflammatory (CD45RChigh) and pro-regulatory (CD45RClow/-) CD8+ T cells in rat, mice and human. 7000 freshly isolated, non-stimulated CD8+ T lymphocytes of four healthy volunteers were analyzed. Combining at a single cell level transcriptome and protein expression data led for the first time to the characterization and definition of three subsets of regulatory CD8+ T cells. Further in vitro functional analysis based on three markers highlighted the superior suppressive activity of the CD8+CD45RClow/-TNFR2+CD29low Tregs subset. To our knowledge, this is the largest characterization of human CD8+ Tregs to date. This data resource will help improve our understanding of CD8+ T cells heterogeneity and will help to translate CD8+ Tregs to the clinic.

immunology↗

bioRxiv2023

XIST dampens X chromosome activity in a SPEN-dependent manner during early human development

XIST long non-coding RNA is responsible for X chromosome inactivation (XCI) in placental mammals, yet it accumulates on both X chromosomes in human female pre-implantation embryos without triggering X chromosome silencing. The long non-coding RNA XACT co-accumulates with XIST on active Xs and may antagonize XIST function. Here we used human ES cells in a naive state of pluripotency to assess the function of XIST and XACT in shaping the X chromosome chromatin and transcriptional landscapes during pre-implantation development. We show that XIST triggers the deposition of polycomb-mediated repressive histone modifications and attenuates transcription of most X-linked genes in a SPEN-dependent manner, while XACT deficiency does not significantly affect XIST activity or X-linked gene expression. Our study demonstrates that XIST is functional prior to XCI, confirms the existence of a transient process of X chromosome dosage compensation, and reveals that X chromosome inactivation and dampening rely on the same set of factors.

molecular biology↗

bioRxiv2023

NINJ1 mediates plasma membrane rupture through formation of nanodisc-like rings

The membrane proteins Ninjurin1 (NINJ1) and Ninjurin2 (NINJ2) are upregulated by nerve injury to increase cell adhesion and promote axonal growth in neurons. NINJ1, but not NINJ2, has also been shown to play an essential role in pyroptosis by promoting plasma membrane rupture downstream of gasdermin D (GSDMD) pore formation, as well as in lytic cell death mediated by other pathways. Recombinant NINJ1 and NINJ2 purified in detergent show irregular rings of various diameters as well as curved filaments. While NINJ1 and NINJ2 both formed ring-like structures when mixed with liposomes, strikingly, only NINJ1, but not NINJ2, ruptures liposome membranes, leading to their dissolution. Because of the better feasibility, we determined the cryo-EM structure of NINJ1 ring segments from detergent by segmenting the irregular rings into shorter fragments. Each NINJ1 subunit contains a transmembrane (TM) helical hairpin (3 and 4) that likely mediates NINJ1 membrane localization, as well as the side-by-side interaction between adjacent subunits. There are two extracellular domain amphipathic helices (1 and 2), among which 1 crosses over to the neighboring subunit at the outside facing surface of the ring, to link NINJ1 subunits together into chains. As such, the inner face of the rings is hydrophobic whereas the outer face of the rings is hydrophilic and should repel membranes. Live cell imaging of NINJ1-deficient THP-1 cells reconstituted with NINJ1-eGFP uncovers the pinching off of NINJ1 rings from the cell surface and the loss of NINJ1 to the culture supernatant in oligomerized forms upon inflammasome activation. Formation of rings is also confirmed by super-resolution imaging of endogenous NINJ1 using anti-NINJ1 antibody. These data suggest that membrane insertion of amphipathic helices and formation of rings with a hydrophilic outer surface underlie the mechanism for NINJ1 to pinch off membranes as if it were a nanodisc-forming amphipathic polymer, leading to membrane rupture and lysis during cell death.

immunology↗

bioRxiv2023

HlyF, an underestimated virulence factor of uropathogenic Escherichia coli

Urinary tract infections (UTIs) are predominantly caused by uropathogenic Escherichia coli (UPEC). By analysing a representative collection of UPEC strains from community-acquired infections, we showed that 20 % of these strains had the ability to produce the protein HlyF. These hlyF+ UPEC strains were the most virulent, mostly responsible for pyelonephritis, often with bloodstream infections. Using a mouse model of UTI, we showed that HlyF was associated with the ability of UPEC to develop a urosepsis, with the presence of bacteria in the spleen and an exacerbated inflammatory response. In contrast to archetypical UPEC strains, hlyF+ UPEC strains are not restricted to phylogroup B2 and harbor a specific repertoire of virulence factors reflecting the fact that HlyF is encoded by conjugative ColV-like plasmids. These plasmids also carry antimicrobial resistance genes, which may facilitate their selection and spreading amongst people receiving antimicrobial therapy. Overall, our data suggest that HlyF is a virulence factor in UPEC and spreading of ColV-like plasmids encoding hlyF warrants further investigation.

microbiology↗