Biology subjects

Paul, J.

Publications and source records attributed to Paul, J..

10 recordsLinked to original sources

bioRxiv2026

Optimized cryo-FIB milling strategy to generate thin, minimally damaged biological lamellae

Focused ion beam (FIB)-milling has been adapted to thin frozen cells for visualization of macromolecular structures in situ with cryogenic electron microscopy. However, only a few large and abundant complexes have been annotated to date. FIB-milling introduces damage which limits the recoverable information from cellular sections. Here, we present Nilas, a low-energy milling strategy optimized to minimize damage and produce thin lamellae. Nilas-milled lamellae show minimal FIB-milling damage, contain areas at or below 50 nm and produce higher resolution in situ 3D reconstructions. Nilas improves the recovery of ribosomal subunits and reduces the predicted minimal detectable molecular mass with two-dimensional template matching (2DTM) to 220 kDa. Consistently, we recover additional non-ribosomal complexes including RNA polymerase III with 2DTM in Nilas-milled lamellae. Nilas is compatible with common milling hardware, making it accessible to diverse users. By extending the size limit for in situ structural biology we bring visual proteomics closer to reality.

biophysics↗

bioRxiv2025

Immunometabolic determinants of long-term response in leukemia patients receiving CD19 CAR T cell therapy

Although most patients with relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL) receiving CD19-targeted chimeric antigen receptor (CAR) T cell therapy achieve remission, loss of CAR T cell functionality and subsequent relapse remains an unmet therapeutic need. We applied an integrative approach to study the immunometabolism of pre- and post-infusion CD19-CAR T cells of patients with relapsed/refractory B-ALL. Pre-infusion CAR T cells of long-term responders (LTR) had increased oxidative phosphorylation, fatty acid oxidation, and pentose phosphate pathway activities, higher mitochondrial mass, tighter cristae, and lower mTOR expression compared to products of short-term responders. Post-infusion CAR T cells in bone marrow (BM) of LTR had high immunometabolic plasticity and mTOR-pS6 expression supported by the BM microenvironment. Transient inhibition of mTOR during manufacture induced metabolic reprogramming and enhanced anti-tumor activity of CAR T cells. Our findings provide insight into immunometabolic determinants of long-term response and suggest a therapeutic strategy to improve long-term remission.

immunology↗

bioRxiv2024

MacqD - A Deep Learning-based Model for Automatic Detection of Socially-housed Laboratory Macaques

Despite advancements in video-based behaviour analysis and detection models for various species, existing methods are suboptimal to detect macaques in complex laboratory environments. To address this gap, we present MacqD, a modified Mask R-CNN model incorporating a SWIN transformer backbone for enhanced attention-based feature extraction. MacqD robustly detects macaques in their home-cage under challenging scenarios, including occlusions, glass reflections, and overexposure to light. To evaluate MacqD and compare its performance against pre-existing macaque detection models, we collected and analysed video frames from 20 caged rhesus macaques at Newcastle University, UK. Our results demonstrate MacqDs superiority, achieving a median F1-score of 99% for frames with a single macaque in the focal cage (surpassing the next-best model by 21%) and 90% for frames with two macaques. Generalisation tests on frames from a different set of macaques from the same animal facility yielded median F1-scores of 95% for frames with a single macaque (surpassing the next-best model by 15%) and 81% for frames with two macaques (surpassing the alternative approach by 39%). Finally, MacqD was applied to videos of paired macaques from another facility and resulted in F1-score of 90%, reflecting its strong generalisation capacity. This study highlights MacqDs effectiveness in accurately detecting macaques across diverse settings.

animal behavior and cognition↗

bioRxiv2024

Direct modulation of CRH nerve terminal function by noradrenaline and corticosterone.

Nerve terminals are the final point of regulation before neurosecretion. As such, neuromodulators acting on nerve terminals can exert significant influence on neural signalling. Hypothalamic corticotropin-releasing hormone (CRH) neurons send axonal projections to the median eminence where CRH is secreted to stimulate the hypothalamic-pituitary-adrenal (HPA) axis. Noradrenaline and corticosterone are two of the most important neuromodulators of HPA axis function; noradrenaline excites CRH neurons and corticosterone inhibits CRH neurons by negative feedback. Here, we used GCaMP6f Ca2+ imaging and measurement of nerve terminal CRH secretion using sniffer cells to determine whether these neuromodulators act directly on CRH nerve terminals. Contrary to expectations, noradrenaline inhibited action potential-dependent Ca2+ elevations in CRH nerve terminals and suppressed evoked CRH secretion. This inhibitory effect was blocked by 2-adrenoreceptor antagonism. Corticosterone also suppressed evoked CRH peptide secretion from nerve terminals, independent of action potential-dependent Ca2+ levels. This inhibition was prevented by the glucocorticoid receptor antagonist, RU486, and indicates that CRH nerve terminals may be a site of fast glucocorticoid negative feedback. Together these findings establish median eminence nerve terminals as a key site for regulation of the HPA axis. SignificanceCorticotropin-releasing hormone (CRH) neurons control the stress axis. Noradrenaline and corticosterone are two signalling molecules that control CRH neuron cell body excitability. However, their effect on CRH nerve terminal function is unknown. To examine this, we performed live Ca2+ imaging and measured CRH secretion. We found that noradrenaline suppressed nerve terminal Ca2+ levels and inhibited nerve terminal CRH secretion. Corticosterone had no effect on nerve terminal Ca2+, but inhibited nerve terminal CRH secretion. This suggests that CRH nerve terminals may be a site of fast corticosteroid negative feedback. Together, these data demonstrate that CRH nerve terminals are a critical point of regulation in the control of the stress axis.

neuroscience↗

bioRxiv2022

Climate stress and its impact on livestock health, farming livelihoods and antibiotic use in Karnataka, India

Understanding the impact of climate change on livestock health is critical to safeguarding global food supplies and economies. Informed by ethnographic research with Indian farmers, veterinarians, and poultry industry representatives, we evidence that both precipitation and vapour pressure are key climate variables that relate to outbreaks of haemorrhagic septicaemia (HS), anthrax (AX), and black quarter (BQ) across the Indian state of Karnataka. We also identify temperature and maximum temperature to be negatively correlated with the same diseases, indicating that a cooling (but still hot) climate with wetter, humid conditions is a prime risk factor for future outbreaks. Principal component analyses have revealed the SW India monsoon and winter periods to be the most strongly correlated with HS, AX and BQ outbreaks. We identify vapour pressure, a proxy for humidity, as having a positive relationship with these specific livestock diseases. The negative relationship between temperature and these diseases, combined with the positive correlation with rainfall and humidity, allow us to classify climate-associated risk using a combination of gridded meteorological time series and epidemiological outbreak data covering the same region and timespan of 1987-2020. Risk maps were constructed following concerns over the growing impact of climate pressures raised by farmers during ethnographic study. Informed by their insights, we used current climate data and future climate projections as a risk classification tool to assess how disease risk varies in Karnataka in the present and possible future scenarios. Despite a relatively limited epidemiological dataset, clear relationships between precipitation, vapour pressure, and temperature with HS, AX and BQ, along with outbreak high-risk zones were defined. This methodology can be replicated to investigate other diseases (including in humans and plants) and other regions, irrespective of scale, as long as the climate and epidemiological data cover similar time periods. This evidence highlights the need for greater consideration of climate change in One Health research and policy and puts forward a case for, we argue, greater alignment between UNFCCC and One Health policy, for example, within the Tripartite Agreement (between OIE, FOA and WHO) on antimicrobial resistance as disease risk cannot be considered independent of climate change. One Health Impact StatementThis research aims to investigate the relationship between factors related to climate (surface temperatures, rainfall, humidity) and outbreaks of livestock-related bacterial diseases. This is especially relevant to the One Health approach as it attempts to integrate findings between not only the science of disease but also the science of climate change as a driver of disease, and address problems that could arise within the public and private sectors (local farming, livestock health, government policy etc.). Providing spatial context to climate-associated disease risk across the Indian state of Karnataka will benefit local farmers that may already be, or transitioning to, more intensive livestock farming along with policy makers and private sector companies who are planning for future investments. This transdisciplinary approach springboards from ethnographic observations of famers lived experiences of challenges to their livelihoods and facilitates the use of climate datasets that may not have been primarily collected for or used by disease-related studies to map long-term epidemiological risk. This demonstrates the pragmatic impact that such transdisciplinary projects can have by providing interpretations of observed risks to animal health (highlighted by social scientists during engagement with practitioner communities) that Earth Scientists were then able to quantify, proving links that would be otherwise not have been evidenced. Using disease data sourced from local institutions, including Government of India facilitates as well academic research laboratories, can plan the application of pragmatic solutions to local farmers who are primarily impacted by the findings of the research.

scientific communication and education↗

bioRxiv2020

Hospital outbreak of carbapenem-resistant Enterobacteriales associated with an OXA-48 plasmid carried mostly by Escherichia coli ST399

A hospital outbreak of carbapenem-resistant Enterobacteriales was detected by routine surveillance. Whole genome sequencing and subsequent analysis revealed a conserved promiscuous OXA-48 carrying plasmid as the defining factor within this outbreak. Four different species of Enterobacteriales were involved in the outbreak. Escherichia coli ST399 accounted for 20/55 of all the isolates. Comparative genomics with publicly available E. coli ST399 sequence data showed that the outbreak isolates formed a unique clade. The OXA-48 plasmid identified in the outbreak differed from other known plasmids by an estimated five homologous recombination events. We estimated a lower bound to the plasmid conjugation rate to be 0.23 conjugation events per lineage per year. Our analysis suggests co-evolution between the plasmid and its main bacterial host to be a key driver of the outbreak. This is the first study to report carbapenem-resistant E. coli ST399 carrying OXA48 as the main cause of a plasmid-borne outbreak within a hospital setting. This study supports complementary roles for both plasmid conjugation and clonal expansion in the emergence of this outbreak.

evolutionary biology↗

bioRxiv2019

Pregnant womens understanding and conceptualisations of the harms from drinking alcohol: a qualitative study

BackgroundDespite womens awareness that drinking alcohol in pregnancy can lead to lifelong disabilities in a child, it appears that an awareness alone does not discourage some pregnant women from drinking. ObjectivesTo explore influences on pregnant womens decision making around alcohol use in a population with frequent and heavy peer drinking (i.e. in two Indigenous Australian communities) and another of non-Indigenous pregnant women attending antenatal care in a range of socioeconomic settings. MethodsIndividual and group discussions were held with both Indigenous Australian and non-Indigenous pregnant women attending a variety of antenatal care models, including two Indigenous maternity services in Australia. Data were analysed using inductive content analysis. ResultsA total of 14 Indigenous Australian and 14 non-Indigenous pregnant women participated in this study. Analysis identified five main influences on pregnant womens alcohol use: the level and detail of womens understanding of harm; womens information sources on alcohol use in pregnancy; how this information influenced their choices; how women conceptualised their pregnancy; and whether the social and cultural environment supported abstinence. Conclusions for practiceThis study provides insight into how Indigenous Australian and non-Indigenous pregnant women understand and conceptualise the harms from drinking alcohol, including how their social and cultural environments impact their ability to abstain. Strategies for behaviour change need to: correct misinformation about supposed safe timing, quantity and types of alcohol; develop a more accurate perception of Fetal Alcohol Spectrum Disorder; reframe messages about harm to messages about optimising the childs health and cognitive outcomes; and develop a holistic approach encompassing womens social and cultural context.

scientific communication and education↗

bioRxiv2018

A Rigorous Interlaboratory Examination of the Need to Confirm NGS-Detected Variants by an Orthogonal Method in Clinical Genetic Testing

Orthogonal confirmation of NGS-detected germline variants has been standard practice, although published studies have suggested that confirmation of the highest quality calls may not always be necessary. The key question is how laboratories can establish criteria that consistently identify those NGS calls that require confirmation. Most prior studies addressing this question have limitations: These studies are generally small, omit statistical justification, and explore limited aspects of the underlying data. The rigorous definition of criteria that separate high-accuracy NGS calls from those that may or may not be true remains a critical issue.\n\nWe analyzed five reference samples and over 80,000 patient specimens from two laboratories. We examined quality metrics for approximately 200,000 NGS calls with orthogonal data, including 1662 false positives. A classification algorithm used these data to identify a battery of criteria that flag 100% of false positives as requiring confirmation (CI lower bound: 98.5-99.8% depending on variant type) while minimizing the number of flagged true positives. These criteria identify false positives that the previously published criteria miss. Sampling analysis showed that smaller datasets resulted in less effective criteria.\n\nOur methodology for determining test and laboratory-specific criteria can be generalized into a practical approach that can be used by many laboratories to help reduce the cost and time burden of confirmation without impacting clinical accuracy.

genetics↗

bioRxiv2017

Copy-number signatures and mutational processes in ovarian carcinoma

Tumours with profound copy-number aberration elude molecular stratification due to their genomic complexity. By representing this complexity as a mixture of copy-number signatures, we provide molecular explanations for differing clinical outcomes. Here we present a method for copy-number signature identification, deriving eight signatures in 117 shallow whole-genome sequenced high-grade serous ovarian cancers (HGSOC), which validated on independent cohorts of 95 deep whole-genome sequenced, and 402 SNP array-profiled cases. Three copy-number signatures predicted longer overall survival, while the others predicted poorer outcome. We found evidence for the mutational processes giving rise to copy-number change for six of the eight signatures via correlations with other genomic features. Our results provide insights into the pathogenesis of HGSOC by uncovering multiple mutational processes that shape genomes following TP53 mutation. Importantly, our work shows that most HGSOC have a mixture of mutational processes suggesting that targeting a single mutator phenotype may be therapeutically suboptimal.

cancer biology↗

bioRxiv2017

Severe infections emerge from the microbiome by adaptive evolution

Bacteria responsible for the greatest global mortality colonize the human microbiome far more frequently than they cause severe infections. Whether mutation and selection within the microbiome accompany infection is unknown. We investigated de novo mutation in 1163 Staphylococcus aureus genomes from 105 infected patients with nose-colonization. We report that 72% of infections emerged from the microbiome, with infecting and nose-colonizing bacteria showing parallel adaptive differences. We found 2.8-to-3.6-fold enrichments of protein-altering variants in genes responding to rsp, which regulates surface antigens and toxicity; agr, which regulates quorum-sensing, toxicity and abscess formation; and host-derived antimicrobial peptides. Adaptive mutations in pathogenesis-associated genes were 3.1-fold enriched in infecting but not nose-colonizing bacteria. None of these signatures were observed in healthy carriers nor at the species-level, suggesting disease-associated, short-term, within-host selection pressures. Our results show that infection, like a cancer of the microbiome, emerges through spontaneous adaptive evolution, raising new possibilities for diagnosis and treatment.\n\nOne Sentence SummaryLife-threatening S. aureus infections emerge from nose microbiome bacteria in association with repeatable adaptive evolution.

genomics↗