Biology subjects

Melo, L.

Publications and source records attributed to Melo, L..

8 recordsLinked to original sources

bioRxiv2026

Age-dependent Effects of Titrating Anodal Transcranial Direct Current Stimulation (tDCS) Intensity on Motor Sequence Learning

Optimising the currently heterogenous efficacy of transcranial direct current stimulation (tDCS) interventions for motor learning requires identifying stimulation parameters facilitating performance while accounting for age-related differences in baseline performance and mechanisms underlying neuroplasticity. We systematically explored anodal tDCS intensity effects on implicit motor sequence learning (IMSL) in young and older adults. The study utilised a randomised, double-blind, counterbalanced crossover design. Ninety-six healthy participants (48 young adults, 48 older adults) completed a serial reaction time task (SRTT) with online sham or anodal tDCS over M1 at intensities of 1, 2, and 3 mA. The next day, memory consolidation was assessed in a recall test. Both age groups demonstrated IMSL and consolidation across conditions. While 1 mA tDCS improved IMSL in young adults by reducing reaction times, higher intensities had no significant benefit compared to sham. In older adults, anodal tDCS did not affect general task performance compared to sham, but 1mA tDCS acutely impaired selective sequence learning. The results demonstrate age-dependent and non-linear dose-dependent effects of anodal tDCS on IMSL. This underscores the necessity for age-adapted protocols for experimental and clinical tDCS applications. Future research should explore neurophysiological reasons for reduced tDCS efficacy in older adults found in the present study.

neuroscience↗

bioRxiv2026

Reusable immobilised quaternary ammonium particles reduce microbial and resistome burdens without promoting resistance selection during wastewater post-treatment.

Wastewater treatment plants act as convergence zones for antibiotic residues, antibiotic-resistant bacteria (ARB), and antimicrobial resistance genes (ARGs), yet conventional processes are not designed to mitigate resistance dissemination from their effluents. While chemical disinfectants are generally effective, soluble quaternary ammonium compounds (QACs) can generate subinhibitory exposure gradients that promote resistance selection and co-selection both during treatment and after release into receiving waters. Here, we evaluate a contact-restricted alternative: benzyldimethyldodecyl ammonium chloride (BDMDAC) immobilised onto hydroxyapatite microparticles as a reusable and retainable post-treatment polishing strategy. Across single-strain assays, treated wastewater exposure, and experimental community evolution, immobilised BDMDAC-functionalised particles (BDMDAC-FPs) achieved concentration-dependent antimicrobial activity without detectable biocide leaching. Optimal exposure (200 mg/L, 4 h) resulted in a ~5.5 log reduction in total bacterial abundance and removal of clinically relevant ARGs. Antimicrobial efficacy was retained after one reuse cycle, supporting operational stability. Plasmid-borne QAC ARGs did not confer protection, and no enrichment of qac-associated or non-QAC ARGs was observed. Conjugation assays demonstrated suppression of horizontal gene transfer even under suboptimal exposure, and mobility-associated markers remained stable or declined during long-term community incubation. Collectively, the data support a contact-restricted mechanism in which antimicrobial pressure is spatially confined to the particle interface, generating high local lethality while limiting diffuse subinhibitory exposure. This spatial confinement decouples antimicrobial efficacy from classical disinfectant-driven resistance selection and mobility amplification. Immobilised BDMDAC-FPs therefore provide a mechanistically distinct and evolution-conscious framework for wastewater polishing technologies.

microbiology↗

bioRxiv2026

Real-time mass-resolved label-free single-molecule immunoassay

Protein-protein interactions govern the biomolecular logic of immunity, signalling, and disease. Elementary rates of association, dissociation, and inhibition underpin our understanding of life processes and remain a limiting factor at the frontier of therapeutic discovery. Yet existing assays infer these processes indirectly through ensemble-averaged signals or molecular labels that obscure native dynamics. Here, we introduce a label-free, real-time, mass-resolved single-molecule immunoassay based on interferometric scattering microscopy (iSCAT) that directly observes individual protein-protein binding events in complex biological samples. By detecting light scattered from single proteins as they bind to an antibody-functionalized surface, we resolve discrete antibody-antigen interactions with single-molecule sensitivity and molecular-weight discrimination. Using IgM as a model system, we demonstrate real-time detection of individual binding events, with measured association rates that scale linearly with concentration over three orders of magnitude. Direct counts of IgM binding events in human serum yield quantitative concentrations that agree with bulk measurements obtained by enzyme-linked immunosorbent assay (ELISA). This bioaffinity iSCAT platform unifies molecular specificity, label-free detection, real-time kinetics, and mass-resolved single-molecule sensitivity, enabling direct access to protein-protein recognition processes and establishing a general framework for quantitative, single-molecule immunoassays.

biophysics↗

bioRxiv2026

Optimizing Network-Level TMS-fMRI: Benchmarking a Novel TMS-Compatible "Sushi" MR Coil

Concurrent TMS-fMRI can map how stimulation affects both the targeted cortex and connected brain-wide networks, but this requires MR receive hardware that allows TMS coil placement while preserving reliable whole-brain BOLD sensitivity. We developed and benchmarked a practical TMS-compatible "Sushi" MR receive setup assembled from two flexible 18-channel body arrays. Across six experiments, we tested functional readout validity, signal quality, and active TMS-fMRI compatibility. Resting-state fMRI (n = 12) and verbal N-back task-fMRI (n = 8) were acquired with Sushi, a commercially available 2x7-channel Surface setup, and a standard 64-channel head/neck array. Functional similarity to the 64-channel reference was quantified with spatial overlap, and multi-echo combination (MEcomb) was tested as a post-acquisition signal optimization strategy. Sushi recovered subject-specific resting-state networks that more closely matched the 64-channel reference than Surface, with no significant difference from the 64-channel test-retest reference. For task-fMRI, MEcomb increased task-map similarity for Sushi, whereas setup comparisons within each pipeline were not significant. MEcomb also improved resting-state similarity and increased temporal signal-to-noise ratio (tSNR) across receive setups. In phantom measurements, TMS coil placement produced spatially graded tSNR reductions relative to the no-TMS-coil condition, strongest near the coil. In one participant, active interleaved single-pulse TMS-fMRI over two cortical sites showed no detectable pulse-locked image artifacts; whole-brain MEcomb tSNR during active TMS-fMRI was reduced by 2.6-6.9% relative to the no-coil/no-stimulation reference. Together, Sushi and MEcomb provide complementary hardware and processing tools for TMS-compatible whole-brain fMRI. This combination supports network-level functional readouts while preserving feasibility for active interleaved TMS-fMRI.

neuroscience↗

bioRxiv2024

The genetic history of Portugal over the past 5,000 years

BackgroundRecent ancient DNA studies uncovering large-scale demographic events in Iberia have focused primarily on Spain, with limited reports for Portugal, a country located at the westernmost edge of continental Eurasia. Here, we introduce the largest collection of ancient Portuguese genomic datasets (n = 68) to date, spanning 5,000 years, from the Neolithic to the 19th century. ResultsWe found evidence of patrilocality in Neolithic Portugal, with admixture from local hunter-gatherers and Anatolian farmers, and persistence of Upper Paleolithic Magdalenian ancestry. This genetic profile persists into the Chalcolithic, reflecting diverse local hunter- gatherer contributions. During the Bronze Age, local genetic ancestry persisted, particularly in southern Iberia, despite influences from the North Pontic Steppe and early Mediterranean contacts. The Roman period highlights Idanha-a-Velha as a hub of migration and interaction, with a notably diverse genetic profile. The Early Medieval period is marked by Central European ancestry linked to Suebi/Visigoth migrations, adding to coeval local, African, and Mediterranean influences. The Islamic and Christian Conquest periods show strong genetic continuity in northern Portugal and significant African admixture in the south, with persistent Jewish and Islamic ancestries suggesting enduring influences in the post-Islamic period. ConclusionsThis study represents the first attempt to reconstruct the genetic history of Portugal from the analysis of ancient individuals. We reveal dynamic patterns of migration and cultural exchange across millennia, but also the persistence of local ancestries. Our findings integrate genetic information with historical and archaeological data, enhancing our understanding of Iberias ancient heritage.

genomics↗

bioRxiv2022

Single-spore germination analyses reveal that Clostridioides difficile uses parallel pathways to sense distinct classes of co-germinants during spore germination.

Clostridioides difficile infections begin when its metabolically dormant spores germinate in response to sensing bile acid germinants alongside amino acid and divalent cation co-germinants in the small intestine. While bile acid germinants are essential for C. difficile spore germination, it is currently unclear whether both co-germinant signals are required. One model proposes that divalent cations, particularly Ca2+, are essential for inducing germination, while another proposes that either co-germinant class can induce germination. The former model is based on the finding that spores defective in releasing large stores of internal Ca2+ in the form of calcium dipicolinic acid (CaDPA) cannot germinate when germination is induced with bile acid germinant and amino acid co-germinant alone. However, since the reduced optical density of CaDPA-less spores makes it difficult to accurately measure their germination, we developed a novel automated, time-lapse microscopy-based germination assay to analyze CaDPA mutant germination at the single-spore level. Using this assay, we found that CaDPA mutant spores germinate in the presence of amino acid co-germinant and bile acid germinant. Higher levels of amino acid co-germinants are nevertheless required to induce CaDPA mutant spores to germinate relative to WT spores because CaDPA released by WT spores during germination can function in a feedforward loop to potentiate the germination of other spores within the population. Collectively, these data indicate that Ca2+ is not essential for inducing C. difficile spore germination because amino acid and Ca2+ co-germinant signals are sensed by parallel signaling pathways. ImportanceClostridioides difficile spore germination is essential for this major nosocomial pathogen to initiate infection. C. difficile spores germinate in response to sensing bile acid germinant signals alongside co-germinant signals. There are two classes of co-germinant signals: Ca2+ and amino acids. Prior work suggested that Ca2+ is essential for C. difficile spore germination based on bulk population analyses of germinating CaDPA mutant spores. Since these assays rely on optical density to measure spore germination and the optical density of CaDPA mutant spores is reduced relative to WT spores, this bulk assay is limited in its capacity to analyze germination. To overcome this limitation, we developed an automated image analysis pipeline to monitor C. difficile spore germination using time-lapse microscopy. With this analysis pipeline, we demonstrate that, although Ca2+ is dispensable for inducing C. difficile spore germination, CaDPA can function in a feedforward loop to potentiate the germination of neighboring spores.

microbiology↗

bioRxiv2022

Ceramide synthase 6 (CerS6) promotes alcohol-induced fatty liver by promoting metabolic dysfunction and upregulating lipid droplet-associated proteins

ObjectiveAlcohol-associated liver disease (ALD) is the leading cause of liver-related mortality worldwide. Current strategies to manage ALD largely focus on advanced stage disease, however, metabolic changes such as glucose intolerance are apparent at the earliest stage of alcoholic steatosis and increase the risk of disease progression. Ceramides impair insulin signaling and accumulate in ALD, and metabolic pathways involving ceramide synthase 6 (CerS6) are perturbed in ALD during hepatic steatosis. In this study, we aimed to investigate the role of CerS6 in ALD development. MethodsC57BL/6 WT and CerS6 KO mice of both sexes were fed either a Lieber-DeCarli control (CON) or 15% ethanol (EtOH) diet for 6 weeks. In vivo metabolic tests including glucose and insulin tolerance tests (GTT and ITT) were performed. The mice were euthanized, and liver histology and lipid levels in serum and liver were measured. For in vitro studies, CerS6 was deleted in human hepatocytes and were incubated with EtOH and/or C16:0-ceramides. RNAseq analysis was performed in mice and in liver from patients with different stages of ALD and diseased controls. ResultsAfter six weeks on an EtOH diet, CerS6 KO mice had reduced body weight, food intake, and %fat mass compared to WT mice. Male (but not female) EtOH-fed KO mice showed significantly higher O2 consumption, CO2 production, respiratory exchange ratio, and energy expenditure (P<0.05 for all) during the dark period compared to EtOH-fed WT mice. In response to EtOH, WT mice developed mild hepatic steatosis, while steatosis was alleviated in KO mice as determined by H&E and ORO staining. KO mice showed significantly decreased long-chain ceramide species, especially C16:0 ceramides, in the serum and liver tissues compared to WT mice. CerS6 deletion decreased serum TG and NEFA only in male not female mice. CerS6 deletion improved glucose tolerance and insulin resistance in EtOH-fed mice of both sexes. RNAseq analysis revealed that 74 genes are significantly upregulated and 66 genes are downregulated by CerS6 deletion in EtOH-fed male mice, with key network pathways including TG biosynthetic process, positive regulation of lipid localization, and fat cell differentiation. Similar to RNAseq results, absence of CerS6 significantly decreased mRNA expression of lipid droplet associated proteins in EtOH-fed mice. In vitro, EtOH stimulation significantly increased PLIN2 protein expression in VL-17A cells while CerS6 deletion inhibited EtOH-mediated PLIN2 upregulation. C16:0-ceramide treatment significantly increased PLIN2 protein expression compared to CON. Importantly, progression of ALD in humans was associated with increased CerS6 hepatic expression. ConclusionsOur findings demonstrate that CerS6 deletion attenuates EtOH-induced weight gain and hepatic steatosis and improves glucose homeostasis in mice fed an EtOH diet. Notably, we unveil that CerS6 plays a major role as a regulator of lipid droplet biogenesis in alcoholic intra-hepatic lipid droplet formation. Together, our data suggest that CerS6 may be targeted for treatment for early stage ALD.

pathology↗

bioRxiv2018

Structure and assembly of a Clostridioides difficile spore polar appendage

Clostridioides difficile, a strict anaerobic spore-former, is the main cause of nosocomial disease associated to antibiotic therapy in adults and a growing concern in the community. Spores are the main infectious, persistence and transmission vehicle. Spore germination occurs in the intestine and the resulting vegetative cells will produce the toxins responsible for the disease symptoms, and spores. During sporulation, a wild type population bifurcates into two main spore morphotypes, with or without a thick exosporium. We show that this bifurcation extends to the formation of spores with a robust polar appendage or spores with a short appendage or that lack this structure. The cysteine-rich CdeM protein localizes to the appendage and around the entire surface of the spore, and is a major structural component of the exosporium, which we show is continuous with the appendage. In a CdeM mutant, when present, the polar appendage is short and disorganized. We show that wild type and cdeM spores with a short or no appendage germinate poorly in response to taurocholate, compared to those with an appendage. cdeM spores of the two types, however, germinate faster than their wild type counterparts. Thus, while the absence of CdeM may increase the permeability of spores to taurocholate, proper assembly of the appendage is also important for germination. Consistent with an overall enhancement of germination, a cdeM mutant shows increased virulence in a hamster model of disease. For a wild type population, spores with a short or no appendage germinate slower than the appendage-bearing spores. Differences in transmission, persistence and disease severity may result, in part, from their proportion in a spore population.

microbiology↗