Biology subjects

Smith, P.

Publications and source records attributed to Smith, P..

At least 19 recordsLinked to original sources

bioRxiv2026

Diagnostic Labels and Measurement Timing Drive Systematic Inconsistency in ADNI Neuroimaging Data

The Alzheimer's Disease Neuroimaging Initiative (ADNI) is widely used to train machine learning models for Alzheimer's disease, yet whether it provides consistent ground truth for predictive modeling has not been systematically tested. In this paper, we showed that ADNI contains three interacting sources of bias with direct implications for machine learning: (a) diagnostic label inconsistency, (b) technical measurement drift, and (c) longitudinal survivor bias. A substantial proportion of cases, particularly within intermediate stages, fall outside ADNI's own diagnostic thresholds. MRI field strength and evolving processing pipelines introduce significant technical variability in hippocampal volume, while cohort survivor bias arising from differential retention of participants across phases further distorts longitudinal estimates of disease progression. These findings indicated that ADNI does not provide the stable, internally consistent labels often required in machine learning applications. We proposed a practical framework for diagnostic validation, feature harmonization, and cohort accounting, offering guidance for building more robust and biologically meaningful predictive models from large-scale neuroimaging cohorts.

neuroscience↗

bioRxiv2026

R-pyocin-mediated selection reverses pan-drug resistance in Pseudomonas aeruginosa

The global escalation of antibiotic resistance is a critical threat necessitating the development of innovative strategies to provide new therapeutic options and restore the efficacy of conventional drugs. Pseudomonas aeruginosa exemplifies this challenge by utilizing a robust genomic resistome to persist in clinical settings. Here, we demonstrate that R-pyocins (phage-like bactericidal particles) can be leveraged not merely as conventional biocides, but as precise selective forces to drive an evolutionary "checkmate" strategy. We subjected the laboratory strains PAO1 and PAK and the clinical pan-drug-resistant (PDR) wound isolate MRSN 6220 to R-pyocin selective pressure. To evade R-pyocins targeting the host lipopolysaccharide (LPS) core, resistance consistently emerges through large-scale chromosomal deletions spanning 250-388 kbp. Crucially, these deletions encompass a conserved region harboring the galU gene (essential for LPS synthesis), the hmgA gene (yielding a pyomelanogenic brown phenotype), and the mexXYZ multidrug efflux operon. While the loss of galU confers broad cross-resistance to R-pyocins by likely truncating the LPS receptor, the concurrent excision of mexXY induces profound collateral sensitivity to aminoglycosides. Furthermore, these large deletions systematically eliminate critical virulence factors and biofilm clusters, including the hcnABC, exoY, phzABCDEFG, and cup operons. In Galleria mellonella and murine chronic wound models, the resulting brown mutants were rendered non-lethal and exhibited a significant 3-log reduction in bacterial load following gentamicin treatment. Ultimately, this work establishes a framework for utilizing R-pyocins as potent evolutionary steering agents to force the predictable reversion of multidrug resistance into an attenuated, biofilm-deficient, and clinically manageable state. Significance StatementPan-drug-resistant (PDR) pathogens demand novel strategies that both kill and restore antibiotic efficacy. Here, we describe an evolutionary checkmate for Pseudomonas aeruginosa, where selection for R-pyocin resistance drives large-scale ([~]300 kb) chromosomal remodeling. Although these deletions confer R-pyocin immunity via loss of the galU gene, they simultaneously collapse the pathogens virulence and defense. Crucially, the excision of the mexXY efflux operon resensitizes PDR strains to conventional aminoglycosides, while the collateral loss of critical virulence and biofilm clusters abrogates pathogenesis. By coupling resistance acquisition to substantial fitness costs, our work establishes a framework for using R-pyocins to force predictable evolutionary trade-offs, driving the reversion of multidrug resistance to an attenuated, biofilm-deficient, and clinically manageable state.

microbiology↗

bioRxiv2026

IL-1β/IL-6 signaling circuit in the tumor microenvironment drives prostate cancer development

Despite considerable progress in elucidating mechanisms leading to castration-resistant prostate cancer (CRPC), insight into the early stages of prostate cancer initiation and progression remains limited. Genomic drivers of prostate cancer initiation have been defined through patient tumor sequencing, but the subsequent events responsible for local tissue invasion are poorly understood. Here we leverage a well-studied genetically engineered mouse prostate cancer model (Hi-Myc) that, based on robust and reproducible kinetics for transitioning from pre-invasive prostatic intraepithelial neoplasia (PIN) to invasive prostate adenocarcinoma (PCa), provides an ideal system to systematically address this question using single-cell analysis. Surprisingly, the transcriptomic profiles of early PIN lesions are indistinguishable from those of late-stage, highly invasive tumor cells, suggesting that MYC activation at the PIN stage establishes a transcriptional program that is fully capable of driving invasion but is restrained by the local tumor microenvironment (TME). Indeed, we find that progression to PCa is associated with progressive infiltration of IL-1{beta}+ tumor-infiltrating macrophages at the PIN stage that, based on immunodepletion and cytokine neutralization experiments, are required for the PIN-to-PCa transition. Mechanistically, IL-1{beta} from macrophages acts directly on prostate fibroblasts, leading to the release of IL-6, which drives invasion by activating IL-6R in tumor cells. Collectively, these findings identify a pro-tumorigenic IL-1{beta}/IL-6 signaling circuit mediated through local macrophages and fibroblasts that unleashes the full oncogenic potential of a cancer driver (MYC) activated at the PIN stage. We also find evidence of this circuit in other (non-MYC-driven) prostate cancer models as well as human prostate and lung adenocarcinoma, with implications for TME-specific targeted therapeutics in early-stage disease.

cancer biology↗

bioRxiv2026

A p53-dependent FBXO44-RAD18 axis limits mutagenesis by terminating translesion DNA synthesis

DNA lesions continually challenge genome replication and threaten genome integrity. DNA damage tolerance pathways, including translesion DNA synthesis (TLS), allow cells to bypass lesions and prevent stalled forks from collapsing into double-strand breaks. Because TLS polymerases are intrinsically error-prone, however, this pathway must be tightly restrained; persistent or deregulated TLS can increase mutagenesis, create therapeutic vulnerabilities, and promote aggressive cancer phenotypes. Through integrated transcriptional profiling, genome-wide CRISPR/Cas9 screening for replication-stress sensitivity, and complementary proteomic analyses, we identify F-box protein 44 (FBXO44) as a late p53-responsive regulator of the TLS mediator RAD18. FBXO44 promotes RAD18 ubiquitination during recovery from replication stress and facilitates shutdown of RAD18-dependent PCNA monoubiquitination. Consistently, FBXO44 loss delays resolution of replication stress and TLS signaling, increases mutation frequency, and is associated with elevated mutational burden and therapy resistance in experimental models and patient datasets. These findings define a p53-FBXO44-RAD18 regulatory axis that limits mutagenic TLS and helps safeguard genome integrity after replication stress.

cancer biology↗

bioRxiv2026

A DMT1-dependent iron-endoplasmic reticulum-extracellular matrix axis regulates cancer cell invasion

Intracellular iron homeostasis is increasingly recognized as a regulator of cancer cell behavior, but how iron distribution influences extracellular matrix (ECM) organization and invasion remains poorly understood. Here, we show that loss of divalent metal transporter 1 (DMT1/SLC11A2) disrupts intracellular iron homeostasis and promotes cancer cell invasion through an iron-ER-ECM axis. In MDA-MB-231 cells, DMT1 knockout (KO) reduced total iron content but increased the labile iron pool (LIP) in both 2D and 3D culture models, indicating altered intracellular iron distribution. Across transcriptomic and phenotypic readouts, DMT1-dependent effects were more evident in 3D than in 2D models, with DMT1 KO inducing endoplasmic reticulum (ER) stress and impaired collagen/ECM organization. Functionally, the DMT1-loss phenotype was marked by reduced 2D motility, whereas in 3D spheroid models DMT1 KO cells displayed enhanced invasive outgrowth in both Matrigel and collagen matrices. Iron chelation further modulated this phenotype in a DMT1-dependent manner. Pharmacologic induction of ER stress phenocopied the loose spheroid architecture and invasive behavior, supporting ER stress as a mechanistic link between altered iron handling and ECM destabilization. Together, these findings identify intracellular iron distribution, rather than total iron abundance alone, as a determinant of ECM integrity and context-dependent cancer cell invasion. One Sentence Summary: Iron misrouting, not iron excess, drives ER stress and collagen failure that unleash invasion in triple-negative breast cancer.

cancer biology↗

bioRxiv2026

Borrelia burgdorferi infection-enhanced Ixodes scapularis serpins S12c5 and S13c5, target MASP1/3 to inhibit MBL-pathway complement activation and promote spirochete survival

Tick serine protease inhibitors (serpins) are critical for tick feeding success and pathogen transmission. We previously demonstrated that two unique serpins, S12c5 and S13c5, are tandemly duplicated on Ixodes scapularis chromosome 5 and are injected into the host at heightened levels during feeding by Borrelia burgdorferi (Bb)-infected nymphs. Here, we characterize the biochemical properties and immunomodulatory functions of these serpins and investigate their roles in promoting Bb transmission. Despite their response to Bb infection, S12c5 and S13c5 share less than 20% overall amino acid identity and differ critically at their reactive center loop (RCL) P1 site, which determines substrate specificity: S13c5 contains a basic arginine residue, while S12c5 contains a polar serine residue. Consistent with these differences, recombinant S13c5 efficiently inhibited innate immune proteases including fXa, plasmin, and trypsin IV (SI: 2.5, 1.9, and 1.0; ka: 8.56 x 103, 3.97 x 10, and 1.08 x 10 M-{superscript 1}s-{superscript 1}, respectively), and consequently delayed plasma clotting via the common pathway, whereas rS12c5 did not inhibit these proteases. However, despite molecular modeling predicting stronger interactions between S13c5 and complement proteases, both serpins inhibited MBL-pathway complement activation by targeting MASP1/3. Deletion of the RCL domain abolished this inhibitory activity, confirming both proteins function as typical inhibitory serpins. Functionally, rS13c5 enhanced Bb colonization in C3H mouse organs. Interestingly, immune response assays revealed a trade-off: S13c5, despite its broader inhibitory activity, was less immunogenic than S12c5. Collectively, these findings establish S12c5 and S13c5 as Bb transmission factors that promote spirochete survival through disruption of host innate immunity.

Molecular Biology↗

bioRxiv2026

The ENIGMA-PD-WML Pipeline: A Containerized, User-Friendly Approach for Accurate, Standardized Segmentation of White Matter Lesions in Multi-Site MRI Data

Understanding vascular contributions to disease is a major unmet need. White matter lesions (WML) are an accepted imaging marker of cerebral small vessel disease, giving insights into its related pathologies. A unified approach for WML analyses in large multi-site data is lacking despite the need for pooling of data to overcome the limitations of often small heterogenous MRI studies which make subtyping and identifying patterns within disease groups difficult. Our ENIGMA-PD-WML pipeline is an open-source containerized pipeline containing all the code and packages required for pre-processing, processing and post-processing of T1-weighted and FLAIR data, outputting accurate and reproducible binary WML maps using a UNet approach. The pipeline provides a standardized image analysis approach for WML and outputs data in both native and MNI space to allow for sharing and pooling of data from multiple sites for large-data analysis. In addition to a reliable standardized approach for WML segmentation, key priorities when developing the pipeline included: usability, i.e., requiring minimal manual input and technical expertise to use, and suitability to run on various MRI scanners and acquisition parameters as is common in multi-site data. This paper describes the pipeline in detail, with rationale for each step, providing transparency and facilitating its usage to overcome reproducibility issues in large-scale WML analyses.

neuroscience↗

bioRxiv2026

Inhibition of the androgen-activating enzyme AKR1C3 selectively decreases systemic and intra-adipose 11-oxygenated androgens in women

Androgen excess drives metabolic and reproductive complications in polycystic ovary syndrome (PCOS), affecting 10-15% of women globally. Aldo-keto reductase 1C3 (AKR1C3) converts inactive precursors from both the classic and the recently identified 11-oxygenated androgen pathways, generating testosterone and 11-ketotestosterone, respectively, which exert comparable androgen receptor activation. Both circulate in similar concentrations in premenopausal women while 11-ketotestosterone is predominant after menopause and in PCOS. Here, we show that adipocytes are a major site of AKR1C3 and androgen receptor expression, with increased expression in women and individuals with obesity. Using human female adipose tissue explants, we find a much higher activation of 11-oxygenated over classic androgens, observing a decrease in 11-oxygenated but not classic androgen activation by AKR1C3 inhibition. Correspondingly, we demonstrate that AKR1C3 inhibitor treatment in premenopausal women selectively disrupts the activation of 11-oxygenated androgens. Pharmacological targeting of AKR1C3 provides a novel strategy to alleviate systemic and intra-adipose 11-oxygenated androgen excess. One Sentence SummaryInhibition of the androgen-activating enzyme AKR1C3 results in a major decrease in 11-oxygenated but not classic androgens in women.

physiology↗

bioRxiv2026

Standalone nanopore sequencing for foodborne pathogen surveillance: a large-scale evaluation and quality control framework

Whole-genome sequencing (WGS) is central to foodborne pathogen surveillance and cross-border outbreak detection. Long-read sequencing using Oxford Nanopore Technologies (ONT) promises rapid, complete, and cost-effective genome assemblies in a single workflow. However, the adoption of standalone ONT sequencing of native DNA has been slowed by concerns that DNA modifications can compromise per-base sequencing accuracy and downstream genotyping. In this study, we evaluated ONT-only sequencing performance across 294 genetically diverse isolates representing ten major foodborne pathogens. Using the SUP@v5.2 basecalling model at 50x coverage, 97.3% (286/294) of the ONT assemblies produced identical or near-identical cgMLST profiles ([≤]3 allelic differences) as Illumina-polished hybrid assemblies. Elevated error rates were observed in four Salmonella enterica serovar Kentucky and four Listeria monocytogenes isolates and were associated with the presence of specific DNA phosphorothioation or methylation systems. Re-basecalling the same dataset with the newly released HAC@v6.0 model revealed a different error profile: although 93.5% (275/294) of assemblies remained highly accurate, all 13 isolates carrying dnd (DNA phosphorothioation) or dpd (7-deazaguanine modification) systems, including isolates of S. enterica, Cronobacter sakazakii, and Vibrio parahaemolyticus, exhibited high error rates, suggesting that such atypical modifications were not adequately represented in the models training dataset. To enable rapid identification of unreliable assemblies, we developed alpaqa, a lightweight computational tool that detects systematic nanopore assembly errors without requiring supplemental short-read data or reference genomes. By identifying affected assemblies, alpaqa provides a quality safeguard for ONT-only workflows. Masking low-quality bases in assemblies flagged by alpaqa improved cgMLST accuracy, although this reduced the number of callable loci and therefore genotyping resolution. Our findings demonstrate that standalone ONT sequencing of native DNA is sufficiently accurate for routine foodborne pathogen surveillance when combined with appropriate quality control, supporting its use in harmonised genomic surveillance frameworks. Data summaryAll sequencing data generated in this study have been submitted to the NCBI Sequence Read Archive. Accession numbers for Illumina and ONT (SUP@v5.2) reads are listed in Supplementary Table S1. Raw pod5 files from error-prone isolates have been deposited in SquiDBase (SQB000021). Alpaqa is available at github.com/MBiggel/alpaqa/. An automated ONT assembly and quality control pipeline integrating alpaqa is available at github.com/MBiggel/boap/. Impact statementThis study demonstrates that standalone Oxford Nanopore sequencing of native DNA can achieve highly accurate genotyping for routine foodborne pathogen surveillance across diverse species. We show that the remaining inaccuracies are linked to specific DNA modification systems, including phosphorothioation and 7-deazaguanine modifications, which are identified here as previously unrecognised sources of systematic sequencing errors. To address this limitation, we introduce alpaqa, a reference-free method for detecting such error-prone assemblies, providing a practical quality-control framework for ONT-only workflows. Together, these results support the reliable use of nanopore sequencing in routine genomic surveillance.

genomics↗

bioRxiv2026

Ruminosignatures associated with methane emissions and feed efficiency across geographies and cattle breeds

The cattle rumen microbiota represents a highly complex and dynamic ecosystem, whose organization and connection to host phenotypes are of the highest importance to food security and the environment. In this study, we analyzed the rumen microbiota, from 2,492 cattle belonging to five different breeds and production systems across five countries, categorizing them into microbial co-abundance groups referred to as Ruminosignatures. We identified twelve distinct Ruminosignatures, including two that were consistently observed across all populations and were dominated by the genus Prevotella and UBA2810. Additional Ruminosignatures showed breed-and diet-specific patterns and collectively explained 96-99% of the variance in rumen microbial composition. The abundances of several Ruminosignatures were associated with methane emissions and feed efficiency, and were influenced by host genetics, with heritability estimates ranging from 0.09 to 0.51. The Ruminosignature dominated by UAB2810 was negatively associated with methane emissions across all datasets and positively linked to feed efficiency in Holstein from Italy and crossbred from Ireland. Additionally, the type of production system affects both the occurrence of Ruminosignatures and their impact on host phenotypes, emphasizing the need for context-specific approaches to modulate the rumen microbiome. Overall, our results offer new perspectives on the assembly of ruminal microbes and underscore the potential of the Ruminosignatures framework for microbiome-informed precision agriculture and breeding initiatives aimed at enhancing feed efficiency and minimizing the environmental impact of cattle farming.

microbiology↗

bioRxiv2025

Identifying win-win opportunities and trade-offs for sustainable agriculture to improve agricultural productivity and soil carbon sequestration: A global meta-analysis

Faced with accelerating climate change and growing food insecurity, sustainable agriculture has the potential to address both issues by enhancing soil carbon sequestration and crop yields. However, it remains unclear whether and when sustainable agricultural practices create win-wins or trade-offs between soil carbon sequestration and food production, and how these outcomes vary across crop types, practice combinations, and environmental conditions. We conducted a global meta-analysis of 2,975 paired yield and topsoil carbon observations (and 498 subsoil carbon observations) from 487 articles to assess the simultaneous impacts of reduced tillage, cover cropping, complex crop rotations, and crop residue retention on soil organic carbon stocks and maize, wheat, rice, and soybean yields. Our analysis revealed that sustainable agricultural practices generally enhanced soil carbon and crop yields, although with considerable heterogeneity. Individual practices often resulted in management trade-offs, as different practices (and crop-practice combinations) optimized either yields or soil carbon sequestration. However, practice stacking often alleviated these trade-offs by combining complementary practices. Win-wins were most likely in marginal agricultural lands characterized by warmer temperatures, water limitation, and low nitrogen application rates - conditions that represent a substantial proportion of global agricultural land area. However, our analysis revealed that the subsoil carbon responses to sustainable agricultural practices were highly variable and may offset topsoil gains, which may limit effective policy design and large-scale implementation.

plant biology↗

bioRxiv2025

Pervasive phenotypic effects of Fbxo42 controlled by regulation of PP4 phosphatase

F-box proteins are the substrate recognition modules of the SCF (SKP1-Cullin- F-box) E3 ubiquitin ligase complex. FBXO42, an understudied member of this family, has recently emerged as a modulator of key cellular processes, including cell cycle progression, DNA damage response and glioma stem cell survival. In this study, we define the function of FBXO42 as a major regulator of the protein phosphatase PP4. Phosphoprotein phosphatases (PPPs) have a broad array of substrates, hence necessitating tight regulation. We observe that FBXO42 ubiquitinates the PP4 complex to govern the assembly of regulatory and catalytic subunits with the net effect of restraining the latters phosphatase activity. FBXO42 depletion unleashes PP4 activity with broad cellular effects highlighting FBXO42 as a novel regulatory node in ubiquitin-mediated signalling for future therapeutic exploitation. Impact StatementFBXO42 is a major regulator of PP4 phosphatase impacting multiple cellular phenotypes.

biochemistry↗

bioRxiv2025

Plasmonic imaging of living pancreatic beta-cell networks

We present a novel plasmonic imaging technique for real-time, label-free tracking of bioelectrical interactions in live pancreatic beta-cell networks. Surface plasmon resonance microscopy (SPRM) is utilized to reveal synchronized glucose-induced intensity oscillations that are suppressed by calcium channel blockers. These oscillations are observed at the subcellular scale with a resolution of 1 m. The technique can also uncover the extracellular spread of these oscillations beyond the cells. We further combine SPRM with network analysis to quantify coordinated electrical activity within the living cell network using both amplitude and phase-based metrics. Our results demonstrate a new method for studying electrical communication in pancreatic beta-cells, which could be crucial for understanding dysregulation in diabetes and advancing treatment development. This technique holds promise for investigating electrical connectivity in biological cell networks with applications in neuroscience, cardiac science, and bioelectricity in cancer, microbiology, development and regeneration. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=114 SRC="FIGDIR/small/649945v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@1dc0ce1org.highwire.dtl.DTLVardef@1d1c333org.highwire.dtl.DTLVardef@6684acorg.highwire.dtl.DTLVardef@906170_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2025

BRCA2 reversion mutation-independent resistance to PARP inhibition in prostate cancer through loss of function perturbations in the DNA pre-replication complex

Recent approvals of PARP inhibitors (PARPi) for BRCA-mutant metastatic castration resistant prostate cancer (mCRPC) necessitate an understanding of the factors that shape sensitivity and resistance. Reversion mutations that restore homologous recombination (HR) repair are detected in [~]50-80% of BRCA-mutant patients who respond but subsequently relapse, but there is currently little insight into why only [~]50% of BRCA-mutant patients display upfront resistance. To address this question, we performed a genome-wide CRISPR screen to identify genomic determinants of PARPi resistance in murine Brca2{Delta}/{Delta} prostate organoids genetically engineered in a manner that precludes the development of reversion mutations. Remarkably, we recovered multiple independent sgRNAs targeting three different members (Cdt1, Cdc6, Dbf4) of the DNA pre-replication complex (pre-RC), each of which independently conferred resistance to olaparib and the next generation PARP-1 selective inhibitor AZD5305. Moreover, sensitivity to PARP inhibition was restored in Brca2{Delta}/{Delta}, Cdc6-depleted prostate cells by knockdown of geminin, a negative regulator of Cdt1, further implicating the critical role of a functional pre-RC complex in PARPi sensitivity. Furthermore, [~]50% of CRPC tumors have copy number loss of pre-RC complex genes, particularly CDT1. Mechanistically, prostate cells with impaired pre-RC activity displayed rapid resolution of olaparib-induced DNA damage as well as protection from replication fork degradation caused by Brca2 loss, providing insight into how Brca2-mutant cancer cells can escape cell death from replication stress induced by PARP inhibition in the absence of HR repair. Of note, a pharmacologic inhibitor that targets the CDT1/geminin complex (AF615) restored sensitivity to AZD5305, providing a potential translational avenue to enhance sensitivity to PARP inhibition in BRCA-mutant cancers. SignificanceHere, we address a major limitation in the effectiveness of PARP inhibitors in BRCA-mutant prostate cancer treatment: only [~]50% of patients respond despite clear genomic evidence of defective homologous recombination. Prior efforts to study PARP inhibitor resistance in prostate cancer have been plagued by the lack of suitable cell lines. We overcame this challenge using primary prostate organoids coupled with genome-wide CRISPR screening. The key finding is that loss of function mutations in the DNA pre-replication complex confer PARP inhibitor resistance. These genes map to chromosomal regions frequently lost in prostate cancer and could therefore serve as potential biomarkers of treatment response. Pharmacologic inhibition of geminin, a negative regulator of the pre-replication complex, can restore PARP inhibitor sensitivity.

cancer biology↗

bioRxiv2025

The Chlamydia trachomatis secreted effector protein CT181 binds to Mcl-1 to prolong neutrophil survival

Chlamydia trachomatis (C.t) infections can lead to severe complications due to the pathogens ability to evade the host immune response, often resulting in asymptomatic infections. The mechanisms underlying this immune subversion remain incompletely understood but likely involve specific bacterial effector proteins. Here, we identify CT181 as a novel effector that directly binds to Mcl-1, a key regulator of neutrophil survival. While a C.t. CT181 mutant exhibited only modest defects in epithelial cell replication and inclusion development, it was essential for C.t. survival in neutrophils, correlating with Mcl-1 stabilization. Using a murine infection model, we demonstrate that CT181 is required for C.t. colonization and cytokine production in vivo. Our findings establish CT181 as the first bacterial effector protein known to bind Mcl-1 to enhance neutrophil survival, revealing a critical strategy by which C.t. promotes immune dysregulation, facilitating bacterial persistence while driving C.t. pathogenesis.

microbiology↗

bioRxiv2025

Partial Wwox Loss of Function Increases Severity of Murine Sepsis and Neuroinflammation

RationaleWW domain-containing oxidoreductase (WWOX) is a gene associated implicated in both neurologic and inflammatory diseases and is susceptible to environmental stressors. We hypothesize partial loss of Wwox function will result in increased sepsis severity and neuroinflammation. MethodsWwox WT/P47T mice, generated by CRISPR/Cas9, and Wwox WT/WT mice were treated with intraperitoneal PBS vs LPS (10mg/kg) and euthanized 12 hours post-injection. Open Field Testing (OFT) and Murine Sepsis Severity Scores (MSS) were utilized to measure sickness behavior and sepsis severity, respectively. Brain tissue was analyzed using immunohistochemistry and PCR to measure neuroinflammation and apoptosis. ResultsWwox WT/P47T LPS mice demonstrated a more significant response to sepsis with an increase in sickness behavior, sepsis severity, gliosis, and apoptosis compared to Wwow WT/WT LPS littermates. ConclusionsPartial loss of Wwox function increases risk for severe sepsis and neuroinflammation. Given the susceptibility of WWOX to environmental stressors, this may be a target for future therapeutic interventions.

neuroscience↗

bioRxiv2024

Genomic Diversity and Antimicrobial Resistance of Vibrio cholerae Isolates from Africa: A PulseNet Africa Initiative Using Nanopore Sequencing to Enhance Genomic Surveillance

ObjectivesVibrio cholerae remains a significant public health threat in Africa, with antimicrobial resistance (AMR) complicating treatment. This study leverages whole-genome sequencing (WGS) of V. cholerae isolates from Cote dIvoire, Ghana, Zambia and South Africa to assess genomic diversity, AMR profiles, and virulence, demonstrating the utility of WGS for enhanced surveillance within the PulseNet Africa network. MethodsWe analysed Vibrio isolates from clinical and environmental sources (2010-2024) using Oxford Nanopore sequencing and hybracter assembly. Phylogenetic analysis, multilocus sequence typing (MLST), virulence and AMR gene detection were performed using Terra, Pathogenwatch, and Cloud Infrastructure for Microbial Bioinformatics (CLMB) platforms, with comparisons against 88 global reference genomes for broader genomic context. ResultsOf 79 high-quality assemblies, 67 were confirmed as V. cholerae, with serogroup O1 accounting for the majority (43/67, 67%). ST69 accounted for 60% (40/67) of isolates, with eight sequence types identified overall. Thirty-seven isolates formed novel sub-clades within AFR12 and AFR15 O1 lineages, suggesting local clonal expansions. AMR gene analysis revealed high resistance to trimethoprim (96%) and quinolones (83%), while resistance to azithromycin, rifampicin, and tetracycline remained low ([≤]7%). A significant proportion of the serogroup O1 isolates (41/43, 95%) harboured resistance genes in at least three antibiotic classes. ConclusionsThis study highlights significant genetic diversity and AMR prevalence in African V. cholerae isolates, with expanding AFR12 and AFR15 clades in the region. The widespread resistance to trimethoprim and quinolones raises concerns for treatment efficacy, although azithromycin and tetracycline remain viable options. WGS enables precise identification of species and genotyping, reinforcing PulseNet Africas pivotal role in advancing genomic surveillance and enabling timely public health responses to cholera outbreaks. Data summaryAll supporting data and protocols have been provided within the article or as supplementary data files. The ONT reads have been deposited under BioProject accession PRJNA1192988, while the high-quality Vibrio spp. assemblies have been shared via figshare (Foster-Nyarko, Ebenezer (2024). Genomic Diversity and Antimicrobial Resistance of Vibrio spp. Isolates from Africa: A PulseNet Africa Initiative Using Nanopore Sequencing to Enhance Genomic Surveillance. figshare. Dataset. https://doi.org/10.6084/m9.figshare.27941376.v1). Individual accession numbers for these reads and Biosample IDs are provided in File S2, available with the online version of this article. The accession numbers for the 88 reference genome assemblies included in our analysis are also provided in File S3. Impact statementCholera remains a significant public health challenge in Africa, disproportionately affecting the region due to the ongoing transmission of Vibrio cholerae O1 and the emergence of antimicrobial resistance (AMR). This study demonstrates the utility of Oxford Nanopore Technology (ONT) sequencing in providing high-resolution insights into the genomic diversity, transmission dynamics, and AMR profiles of V. cholerae isolates across Africa. By generating and analysing whole-genome sequences, we identified novel sublineages, high prevalence rates of AMR genes, and virulence traits critical to cholera pathogenesis. These findings contribute to a deeper understanding of the epidemiology and evolution of V. cholerae in Africa, informing targeted intervention strategies. Furthermore, the study highlights the growing threat posed by AMR among V. cholerae isolates, including resistance to key therapeutic antibiotics, such as quinolones and trimethoprim, which could undermine current treatment protocols. Despite this, the absence of resistance to azithromycin and rifampicin among the O1 isolates suggests these drugs may remain viable treatment options, offering a critical avenue for preserving treatment efficacy. This research also underscores the importance of sustained genomic surveillance, capacity building, and regional collaboration to mitigate the public health impact of cholera and other foodborne pathogens. By leveraging WGS technologies and training initiatives, such as the PulseNet Africa genomics workshop, this study provides a framework for strengthening regional capacities to detect, monitor, and respond to cholera outbreaks and the spread of AMR. These efforts align with the African Union and Africa CDCs strategic priorities on health security and AMR, contributing to improved public health systems and cholera control across the continent.

genomics↗

bioRxiv2024

Tetramer formation of CpoS facilitates Inc-Inc interactions during Chlamydia trachomatis infection

Chlamydia trachomatis (C.t.), the leading bacterial cause of sexually transmitted infections, replicates within a unique intracellular compartment called the inclusion, which is modified by secreted proteins known as inclusion membrane (Inc) proteins. Here we further characterize CpoS, an Inc previously shown to be critical for replication and inclusion development. We demonstrate that CpoS directly binds multiple coiled-coil domain-containing Incs while simultaneously engaging Rab GTPases at a separate site. Notably, CpoS-InaC interactions facilitate the recruitment of select Arfs to the inclusion membrane, while Rab recruitment occurs independtly of these interactions. Biochemical and biophysical analyses revealed that Incs self-oligomerize, forming higher-ordered structures, with CpoS adpoting a tetrameric structure resembling eukaryotic SNAREs. We propose these assemblies likely serve as scaffolds to orchestrate vesicle docking, tethering, and fusion. Our findings underscore the intricate interplay between bacterial and host factors, revealing that C.t. leverages both Inc-Inc interactions and host protein engagement to manipulate vesicular trafficking and sustain infection.

microbiology↗