Biology subjects

Douglas, A.

Publications and source records attributed to Douglas, A..

10 recordsLinked to original sources

bioRxiv2026

A stable, hierarchical LIN code system for Campylobacter jejuni and Campylobacter coli: A unified genomic nomenclature for lineage-level typing and global surveillance.

Campylobacter remains the leading cause of bacterial gastroenteritis worldwide, with C. jejuni accounting for around 90% of infection and C. coli accounting for most of the rest. Seven-locus multilocus sequence typing (MLST) has improved our understanding of host association and population structure, whilst core genome MLST (cgMLST), enables investigation of transmission events at high-resolution. However, the lack of a stable and standardised nomenclature for clustering of cgMLST data has limited reproducibility and long-term comparability between studies. Here we introduce a joint, hierarchical Life Identification Number (LIN) code system that provides reproducible, multi-level genomic identifiers for C. jejuni and C. coli lineages. Using an updated cgMLST v2 scheme (1,142 loci) and globally representative datasets of high-quality genomes selected from over 53,000 assemblies in the Campylobacter PubMLST database (https://pubmlst.org/organisms/campylobacter-jejunicoli), we firstly defined LIN codes on a dataset of 5,664 genomes. Pairwise allelic distances were computed using MSTclust, and 18 nested thresholds were defined through silhouette, adjusted Wallace and adjusted Rand Index (ARI) statistics to capture the population structure from species to outbreak level resolution. The LIN thresholds were then validated using a second dataset of 1,781 genomes from PubMLST and applied to a large water-associated outbreak dataset from New Zealand in 2016, containing clinical and ecological genomes. Further application of LIN codes was demonstrated by analyses of the C. jejuni ST-21 clonal complex and ST-6175 isolates, as well as the broader population structure of C. coli, using data from PubMLST. Across all datasets, LIN clusters were stable, largely monophyletic, and back-compatible with existing nomenclature, accurately distinguishing host-adapted and outbreak-associated lineages. By embedding cgMLST data within a stable and scalable nomenclature, the Campylobacter LIN system delivers consistent, automated genome-to-lineage assignment. This unified framework bridges population genetics and applied surveillance, enabling robust, real-time comparison of Campylobacter isolates across sources, studies, and time. Impact statementHuman cases of Campylobacter worldwide continue unabated. Tracing the source of Campylobacter infection is particularly challenging given the sporadic or multi-source nature of outbreaks, with potential transmission from foodborne, animal or environmental sources. Seven-locus MLST has greatly improved our broad understanding of Campylobacter population structure. However, whilst high-resolution cgMLST alleles and STs themselves do not change, longitudinal cluster analyses of cgMLST data have lacked a stable nomenclature, rendering them unsuitable for robust and comparable surveillance over time. Life Identification Number (LIN) codes provide a solution to this problem, establishing an automated and scalable nomenclature derived directly from cgMLST profiles, that is stable over time. We have implemented a joint C. jejuni and C. coli LIN code scheme in PubMLST, with scripts for real-time lineage assignment. LIN codes are back-compatible with existing MLST nomenclature, and we demonstrate their added practical value for exploring population structure and high-resolution outbreak investigation. LIN codes support surveillance of Campylobacter in a One Health context, by enabling consistent typing at multiple levels across different sources, laboratories and time. Data summary1. The isolate collections used to develop the LIN codes are publicly available and searchable as individual projects on the PubMLST database (https://pubmlst.org). O_LILIN code development (Dataset 1) (n=5,664 isolates, up to 200 isolates per clonal complex) C_LIO_LILIN code validation (Dataset 2) (n=1,781 isolates), up to 50 isolates per clonal complex C_LIO_LIOutbreak investigation (Dataset 3): New Zealand 2016 Havelock North waterborne outbreak, Gilpin et al (n=161 isolates) [1] C_LIO_LIPopulation structure exploration (clonal complex) (Dataset 4): ST-21 complex (n=1800 isolates, up to 100 isolates randomly selected from each country) C_LIO_LIPopulation structure exploration (sequence type) (Dataset 5): ST-6175 (n=321 isolates, genomes with good cgMLST v2 annotation) C_LI 2. The software for LIN code development is publicly available as follows: O_LIMSTclust for pairwise distance matrices https://gitlab.pasteur.fr/GIPhy/MSTclust [2] C_LIO_LIPython script to define LIN codes in a local dataset; (https://gitlab.pasteur.fr/BEBP/LINcoding) C_LIO_LIBIGSdb Perl script to define LIN codes from cgMLST profiles on the PubMLST database; (https://github.com/kjolley/BIGSdb/blob/develop/scripts/maintenance/lincodes.pl) C_LI

microbiology↗

bioRxiv2025

Sickness engrams modulate anticipatory immune responses

A threat to survival in the wild is vulnerability to infection. The immune system is essential for defence against foreign species which cause sickness. During infection the brain triggers conserved behaviors, including fever, tiredness and anorexia that support immunity. The immune system stores infection information via adaptive immunity, however it remains unclear whether the brain stores immune-related information as long-term memory engrams. Here we demonstrate that mice form contextual memories for sickness events. Upon sickness-memory recall mice lower whole-body metabolism, and increase coactivation between the hippocampus and sickness regions such as the central amygdala, alongside elevated engram activation. Optogenetic reactivation of sickness engrams decreases metabolism, similar to natural recall. Finally, natural recall and artificial reactivation of a sickness-memory increased genes associated with the acute phase response in the liver. These findings suggest that sickness experiences are encoded as engrams, which upon reactivation trigger coordinated metabolic and innate immune responses.

neuroscience↗

bioRxiv2025

YAP1 and QSER1 are Key Modulators of Embryonic Signaling Pathways in the Mammalian Epiblast

YAP1 signaling is essential for development but its specific roles in early embryogenesis remain poorly understood. To shed light on this, we analyzed YAP1s role in regulating the pluripotency of the mammalian epiblast, using scRNAseq approaches. Conditional deletion of Yap1 in the mouse epiblast (Sox2-Cre) altered the expression of signaling genes, including Nodal, Wnt3, and Fgf8. Accordingly, Yap1 loss led to enhanced differentiation of the epiblast toward primitive streak lineages, as evidenced by the upregulation of T/Brachyury and Eomes genes. Furthermore, a proximity labeling assay in human pluripotent stem cells, followed by biochemical assays and molecular modeling predictions, revealed that YAP1 cooperates with QSER1 protein to regulate lineage genes. Our analysis shows that YAP1:TEAD4 enhancers recruit QSER1 to prevent RNA Polymerase II recruitment. Accordingly, QSER1 depletion, similar to YAP1, increases NODAL gene expression and leads to hyperactive NODAL signaling in human 2D-gastruloids. Overall, our findings define a role of YAP1 in the epiblast in vivo and uncovered an interplay with QSER1 controlling the activity of developmental signaling pathways in pluripotent cells.

developmental biology↗

bioRxiv2021

Accelerating manufacturing to start large-scale supply of a new adenovirus-vectored vaccine within 100 days

The Coalition for Epidemic Preparedness Innovations 100-day moonshot aspires to launch a new vaccine within 100 days of pathogen identification. Here, we describe work to optimize adenovirus vector manufacturing for rapid response, by minimizing time to clinical trial and first large-scale supply, and maximizing the output from the available manufacturing footprint. We describe a rapid viral seed expansion workflow that allows vaccine release to clinical trials within 60 days of antigen sequence identification, followed by vaccine release from globally distributed sites within a further 40 days. We also describe a new perfusion-based upstream production process, designed to maximize output while retaining simplicity and suitability for existing manufacturing facilities. This improves upstream volumetric productivity of ChAdOx1 nCoV-19 by around four-fold and remains compatible with the existing downstream process, yielding drug substance sufficient for 10000 doses from each liter of bioreactor capacity. Transition to a new production process across a large manufacturing network is a major task. In the short term, the rapid seed generation workflow could be used with the existing production process. We also use techno-economic modelling to show that, if linear scale-up were achieved, a single cleanroom containing two 2000 L bioreactors running our new perfusion-based process could supply bulk drug substance for around 120 million doses each month, costing <0.20 EUR/dose. We estimate that a manufacturing network with 32000 L of bioreactor capacity could release around 1 billion doses of a new vaccine within 130 days of genomic sequencing of a new pathogen, in a hypothetical surge campaign with suitable prior preparation and resources, including adequate fill-and-finish capacity. This accelerated manufacturing process, along with other advantages such as thermal stability, supports the ongoing value of adenovirus-vectored vaccines as a rapidly adaptable and deployable platform for emergency response.

bioengineering↗

bioRxiv2021

The Predicted Metabolic Function of the Gut Microbiota of Drosophila melanogaster

An important goal for many nutrition-based microbiome studies is to identify the metabolic function of microbes in complex microbial communities and its impact on host physiology. This research can be confounded by poorly-understood effects of community composition and host diet on the metabolic traits of individual taxa. Here, we investigated these multi-way interactions by constructing and analyzing metabolic models comprising every combination of five bacterial members of the Drosophila gut microbiome (from single taxa to the five-member community of Acetobacter and Lactobacillus species) under three nutrient regimes. We show that the metabolic function of Drosophila gut bacteria is dynamic, influenced by community composition and responsive to dietary modulation. Furthermore, we show that ecological interactions such as competition and mutualism identified from the growth patterns of gut bacteria are underlain by a diversity of metabolic interactions, and show that the bacteria tend to compete for amino acids and B vitamins more frequently than for carbon sources. Our results reveal that in addition to fermentation products such as acetate, intermediates of the tricarboxylic acid (TCA) cycle including 2-oxoglutarate and succinate are produced at high flux and cross-fed between bacterial taxa suggesting important roles for TCA cycle intermediates in modulating Drosophila gut microbe interactions and the potential to influence host traits. These metabolic models provide specific predictions of the patterns of ecological and metabolic interactions among gut bacteria under different nutrient regimes, with potentially important consequences for overall community metabolic function and nutritional interactions with the host. IMPORTANCEDrosophila is an important model for microbiome research partly because of the low complexity of its mostly culturable gut microbiota. Our current understanding of how Drosophila interacts with its gut microbes and how these interactions influence host traits derives almost entirely from empirical studies that focus on individual microbial taxa or classes of metabolites. These studies have failed to capture fully the complexity of metabolic interactions that occur between host and microbe. To overcome this limitation, we reconstructed and analyzed 31 metabolic models for every combination of the five principal bacterial taxa in the gut microbiome of Drosophila. This revealed that metabolic interactions between between Drosophila gut bacterial taxa are highly dynamic and influenced by co-occurring bacteria and nutrient availability. Our results generate testable hypothesis about among-microbe ecological interactions in the Drosophila gut and the diversity of metabolites available to influence host traits.

microbiology↗

bioRxiv2020

Full species-wide leaf and seed ionomic diversity of Arabidopsis thaliana

O_LISoil is a heterogenous reservoir of essential elements needed for plant growth and development. Plants have evolved mechanisms to balance their nutritional needs based on availability of nutrients. This has led to genetically-based variation in the elemental composition ionome, of plants, both within and between species. C_LIO_LIWe explore this natural variation using a panel of wild-collected, geographically widespread Arabidopsis thaliana accessions from the 1001 Genomes Project including over 1,135 accessions, and the 19 parental accessions of the Multi-parent Advanced Generation Inter-Cross (MAGIC) panel, all with full-genome sequences available. C_LIO_LIWe present an experimental design pipeline for high-throughput ionomic screenings and analyses with improved normalisation procedures to account for errors and variability in conditions often encountered in large-scale, high-throughput data collection. We report quantification of the complete leaf and seed ionome of the entire collection using this pipeline and a digital tool-IonExplorer to interact with the dataset. C_LIO_LIWe describe the pattern of natural ionomic variation across the A. thaliana species and identify several accessions with extreme ionomic profiles. It forms a valuable resource for exploratory QTL, GWA studies to identify genes underlying natural variation in leaf and seed ionome and genetic adaptation of plants to soil conditions. C_LI

plant biology↗

bioRxiv2019

Correction of amyotrophic lateral sclerosis related phenotypes in induced pluripotent stem cell-derived motor neurons carrying a hexanucleotide expansion mutation in C9orf72 by CRISPR/Cas9 genome editing using homology-directed repair

The G4C2 hexanucleotide repeat expansion (HRE) in C9orf72 is the commonest cause of familial amyotrophic lateral sclerosis (ALS). A number of different methods have been used to generate isogenic control lines using CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 and non-homologous end-joining (NHEJ) by deleting the repeat region with the risk of creating indels and genomic instability. In this study we demonstrate complete correction of an induced pluripotent stem cell (iPSC) line derived from a C9orf72-HRE positive ALS/FTD patient using CRISPR/Cas9 genome editing and homology directed repair (HDR), resulting in replacement of the excised region with a donor template carrying the wild-type repeat size to maintain the genetic architecture of the locus. The isogenic correction of the C9orf72 HRE restored normal expression and methylation at the C9orf72 locus, reduced intron retention in the edited lines, and abolished pathological phenotypes associated with the C9orf72 HRE expansion in iPSC derived motor neurons (iPSMNs). RNA sequencing of the mutant line identified 2220 differentially expressed genes compared to its isogenic control. Enrichment analysis demonstrated an over-representation of ALS relevant pathways, including calcium ion dependent exocytosis, synaptic transport and the KEGG ALS pathway, as well as new targets of potential relevance to ALS pathophysiology. Complete correction of the C9orf72 HRE in iPSMNs by CRISPR/Cas9 mediated HDR provides an ideal model to study the earliest effects of the hexanucleotide expansion on cellular homeostasis and the key pathways implicated in ALS pathophysiology.

neuroscience↗

bioRxiv2019

Adaptations of energy metabolism in cetaceans have consequences for their response to foraging disruption

Cetaceans have varied their anatomical structure, physiology and metabolism to adapt to the challenges of aquatic life. Key to this change is the deposition of blubber. This adipose tissue plays a significant regulatory and signaling role in mammalian metabolism. As foraging disruption by human activities is emerging as a key conservation threat for cetaceans, we need to understand how selection for aquatic life might have altered key nutrient sensing pathways associated with adipose signaling. We compared selection pressure on those energy metabolism biological pathways by contrasting the rate of substitution observed in genes associated with them in cetacean and artiodactyl genomes. We then estimated the likely consequence of these selection pressures for pathway functions. Here we show that genes involved in the insulin, mTOR, SIRT and NF-{kappa}B pathways were under significant positive selection in cetaceans compared to their terrestrial sister taxon. Our results suggest these genes may have been positively selected to adapt to a glucose-poor diet and it is unlikely that fat depots signaling function in the same manner as in terrestrial mammals. Secondary adaptation to life in water significantly affected functions in nutrient sensing pathways in cetaceans. Insulin is not likely to play the same role in energy balance as it does in terrestrial mammals and adiposity is not likely to have the deleterious health consequences it has in terrestrial mammals. The physiological ecology of cetacean fat deposition, and therefore its value as a condition index, needs to be interpreted in this evolutionary context.

evolutionary biology↗

bioRxiv2018

A simian-adenovirus-vectored rabies vaccine suitable for thermostabilisation and clinical development for low-cost single-dose pre-exposure prophylaxis

BackgroundEstimates of current global rabies mortality range from 26,000 to 59,000 deaths per annum. Although pre-exposure prophylaxis using inactivated rabies virus vaccines (IRVs) is effective, it requires two to three doses and is regarded as being too expensive and impractical for inclusion in routine childhood immunization programmes.\n\nMethodology/ Principal FindingsHere we report the development of a simian-adenovirus-vectored rabies vaccine intended to enable cost-effective population-wide pre-exposure prophylaxis against rabies. ChAdOx2 RabG uses the chimpanzee adenovirus serotype 68 (AdC68) backbone previously shown to achieve pre-exposure protection against rabies in non-human primates. ChAdOx2 differs from AdC68 in that it contains the human adenovirus serotype 5 (AdHu5) E4 orf6/7 region in place of the AdC68 equivalents, enhancing ease of manufacturing in cell lines which provide AdHu5 E1 proteins in trans.\n\nWe show that immunogenicity of ChAdOx2 RabG in mice is comparable to that of AdC68 RabG and other adenovirus serotypes expressing rabies virus glycoprotein. High titers of rabies virus neutralizing antibody (VNA) are elicited after a single dose. The relationship between levels of VNA activity and rabies glycoprotein monomer-binding antibody differs after immunization with adenovirus-vectored vaccines and IRV vaccines, suggesting routes to further enhancement of the efficacy of the adenovirus-vectored candidates. We also demonstrate that ChAdOx2 RabG can be thermostabilised using a low-cost method suitable for clinical bio-manufacture and ambient-temperature distribution in tropical climates. Finally, we show that a dose-sparing effect can be achieved by formulating ChAdOx2 RabG with a simple chemical adjuvant. This approach could lower the cost of ChAdOx2 RabG and other adenovirus-vectored vaccines.\n\nConclusions/ SignificanceChAdOx2 RabG may prove to be a useful tool to reduce the human rabies death toll. We have secured funding for Good Manufacturing Practice-compliant bio-manufacture and Phase I clinical trial of this candidate.\n\nAuthor summaryRabies was, after smallpox, the second human disease for which an efficacious vaccine was developed, by Pasteur in 1885. Although it is eminently preventable, with highly efficacious vaccines available for both humans and animals, it still causes considerable mortality in low and middle-income countries. It is a particular problem in areas with the weakest healthcare and veterinary infrastructure, where achieving prompt post-exposure vaccination or high-coverage dog vaccination are challenging.\n\nHere, we report the development of a new candidate rabies vaccine, designed to enable low-cost single-dose pre-exposure human rabies prophylaxis in such settings. ChAdOx2 RabG is based upon a simian adenovirus-vectored candidate previously shown to achieve protection after a single dose in non-human primates, now modified to allow clinical-grade bio-manufacture. We show that it induces a potent immune response in mice, that this response can be further enhanced by clinically-relevant adjuvant, and that we can stabilise it such that it can withstand temperatures of up to 45 {degrees}C for a month. We will be performing a clinical trial of this candidate in the near future.

immunology↗

bioRxiv2018

Genome-wide analysis in UK Biobank identifies over 100 QTLs associated with muscle mass variability in middle age individuals

Muscle bulk in adult healthy humans is highly variable even after accounting for height, age and sex. Low muscle mass, due to fewer and/or smaller constituent muscle fibers, would exacerbate the impact of muscle loss occurring in aging or disease. Genetic variability substantially influences muscle mass differences, but causative genes remain largely unknown. In a genome-wide association study (GWAS) on appendicular lean mass (ALM) in a population of 85,750 middle-age (38-49 years) individuals from the UK Biobank (UKB) we found 182 loci associated with ALM (P<5x10-8). We replicated associations for 78% of these loci (P<5x10-8) with ALM in a population of 181,862 elderly (60-74 years) individuals from UKB. We also conducted a GWAS on hindlimb skeletal muscle mass of 1,867 mice from an advanced intercross between two inbred strains (LG/J and SM/J) which identified 23 quantitative trait loci. 38 positional candidates distributed across 5 loci overlapped between the two species. In vitro studies of positional candidates confirmed CPNE1 and STC2 as modifiers of myogenesis. Collectively, these findings shed light on the genetics of muscle mass variability in humans and identify targets for the development of interventions for treatment of muscle loss. The overlapping results between humans and the mouse model GWAS point to shared genetic mechanisms across species.

genetics↗