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An Interdisciplinary Research Centre at the University of Cambridge
 

Genetic diversity, determinants, and dissemination of <em>Burkholderia pseudomallei</em> lineages implicated in melioidosis in northeast Thailand

Mon, 18/12/2023 - 11:00

bioRxiv. 2023 Dec 10:2023.06.02.543359. doi: 10.1101/2023.06.02.543359. Preprint.

ABSTRACT

Melioidosis is an often-fatal neglected tropical disease caused by an environmental bacterium Burkholderia pseudomallei. However, our understanding of the disease-causing bacterial lineages, their dissemination, and adaptive mechanisms remains limited. To address this, we conducted a comprehensive genomic analysis of 1,391 B. pseudomallei isolates collected from nine hospitals in northeast Thailand between 2015 and 2018, and contemporaneous isolates from neighbouring countries, representing the most densely sampled collection to date. Our study identified three dominant lineages with unique gene sets enhancing bacterial fitness, indicating lineage-specific adaptation strategies. Crucially, recombination was found to drive lineage-specific gene flow. Transcriptome analyses of representative clinical isolates from each dominant lineage revealed heightened expression of lineage-specific genes in environmental versus infection conditions, notably under nutrient depletion, highlighting environmental persistence as a key factor in the success of dominant lineages. The study also revealed the role of environmental factors - slope of terrain, altitude, direction of rivers, and the northeast monsoons - in shaping B. pseudomallei geographical dispersal. Collectively, our findings highlight persistence in the environment as a pivotal element facilitating B. pseudomallei spread, and as a prelude to exposure and infection, thereby providing useful insights for informing melioidosis prevention and control strategies.

PMID:38106061 | PMC:PMC10723255 | DOI:10.1101/2023.06.02.543359

Prenatal cortisol exposure impairs adrenal function but not glucose metabolism in adult sheep

Mon, 18/12/2023 - 11:00

J Endocrinol. 2023 Dec 1:JOE-23-0326. doi: 10.1530/JOE-23-0326. Online ahead of print.

ABSTRACT

Adverse environmental conditions before birth are known to program adult metabolic and endocrine phenotype in several species. However, whether increments in fetal cortisol concentrations of the magnitude commonly seen in these conditions can cause developmental programming remains unknown. Thus, this study investigated the outcome of physiological increases in fetal cortisol concentrations on glucose-insulin dynamics and pituitary-adrenal function in adult sheep. Compared to saline treatment, intravenous fetal cortisol infusion for 5 days in late gestation did not affect birthweight but increased lamb body weight at 1-2 weeks after birth. Adult glucose dynamics, insulin sensitivity and insulin secretion were unaffected by prenatal cortisol overexposure, assessed by glucose tolerance tests, hyperinsulinaemic-euglycaemic clamps and acute insulin administration. In contrast, prenatal cortisol infusion induced adrenal hypo-responsiveness in adulthood with significantly reduced cortisol responses to insulin-induced hypoglycaemia and exogenous adrenocorticotropic hormone (ACTH) administration relative to saline treatment. The area of adrenal cortex expressed as a percentage of the total cross-sectional area of the adult adrenal gland was also lower after prenatal cortisol than saline infusion. In adulthood, basal circulating ACTH but not cortisol concentrations were significantly higher in the cortisol than saline treated group. The results show that cortisol overexposure before birth programs pituitary-adrenal development with consequences for adult stress responses. Physiological variations in cortisol concentrations before birth may, therefore, have an important role in determining adult phenotypical diversity and adaptability to environmental challenges.

PMID:38109257 | DOI:10.1530/JOE-23-0326

Site-selective peptide functionalisation mediated <em>via</em> vinyl-triazine linchpins

Mon, 18/12/2023 - 11:00

Chem Commun (Camb). 2023 Dec 18. doi: 10.1039/d3cc05213c. Online ahead of print.

ABSTRACT

Herein we introduce 3-vinyl-1,2,4-triazines derivatives as dual-reactive linkers that exhibit selectivity towards cysteine and specific strained alkynes, enabling conjugate addition and inverse electron-demand Diels-Alder (IEDDA) reactions. This approach facilitates site-selective bioconjugation of biologically relevant peptides, followed by rapid and highly selective reactions with bicyclononyne (BCN) reagents.

PMID:38108130 | DOI:10.1039/d3cc05213c

Acquisition of suppressive function by conventional T cells limits antitumor immunity upon T<sub>reg</sub> depletion

Fri, 15/12/2023 - 11:00

Sci Immunol. 2023 Dec 15;8(90):eabo5558. doi: 10.1126/sciimmunol.abo5558. Epub 2023 Dec 15.

ABSTRACT

Regulatory T (Treg) cells contribute to immune homeostasis but suppress immune responses to cancer. Strategies to disrupt Treg cell-mediated cancer immunosuppression have been met with limited clinical success, but the underlying mechanisms for treatment failure are poorly understood. By modeling Treg cell-targeted immunotherapy in mice, we find that CD4+ Foxp3- conventional T (Tconv) cells acquire suppressive function upon depletion of Foxp3+ Treg cells, limiting therapeutic efficacy. Foxp3- Tconv cells within tumors adopt a Treg cell-like transcriptional profile upon ablation of Treg cells and acquire the ability to suppress T cell activation and proliferation ex vivo. Suppressive activity is enriched among CD4+ Tconv cells marked by expression of C-C motif receptor 8 (CCR8), which are found in mouse and human tumors. Upon Treg cell depletion, CCR8+ Tconv cells undergo systemic and intratumoral activation and expansion, and mediate IL-10-dependent suppression of antitumor immunity. Consequently, conditional deletion of Il10 within T cells augments antitumor immunity upon Treg cell depletion in mice, and antibody blockade of IL-10 signaling synergizes with Treg cell depletion to overcome treatment resistance. These findings reveal a secondary layer of immunosuppression by Tconv cells released upon therapeutic Treg cell depletion and suggest that broader consideration of suppressive function within the T cell lineage is required for development of effective Treg cell-targeted therapies.

PMID:38100544 | DOI:10.1126/sciimmunol.abo5558

A recombinant approach for stapled peptide discovery yields inhibitors of the RAD51 recombinase

Mon, 11/12/2023 - 11:00

Chem Sci. 2023 Nov 21;14(47):13915-13923. doi: 10.1039/d3sc03331g. eCollection 2023 Dec 6.

ABSTRACT

Stapling is a macrocyclisation method that connects amino acid side chains of a peptide to improve its pharmacological properties. We describe an approach for stapled peptide preparation and biochemical evaluation that combines recombinant expression of fusion constructs of target peptides and cysteine-reactive divinyl-heteroaryl chemistry as an alternative to solid-phase synthesis. We then employ this workflow to prepare and evaluate BRC-repeat-derived inhibitors of the RAD51 recombinase, showing that a diverse range of secondary structure elements in the BRC repeat can be stapled without compromising binding and function. Using X-ray crystallography, we elucidate the atomic-level features of the staple moieties. We then demonstrate that BRC-repeat-derived stapled peptides can disrupt RAD51 function in cells following ionising radiation treatment.

PMID:38075664 | PMC:PMC10699557 | DOI:10.1039/d3sc03331g

Recapitulating primary immunodeficiencies with expanded potential stem cells: Proof of concept with STAT1 gain of function

Sun, 10/12/2023 - 11:00

J Allergy Clin Immunol. 2023 Dec 9:S0091-6749(23)02411-9. doi: 10.1016/j.jaci.2023.11.914. Online ahead of print.

ABSTRACT

BACKGROUND: Inborn errors of immunity (IEI) often lack specific disease models and personalized management. Signal transducer and activator of transcription (STAT)-1 gain of function (GoF) is such example of an IEI with diverse clinical phenotype with unclear pathomechanisms and unpredictable response to therapy. Limitations in obtaining fresh samples for functional testing and research further highlights the need for patient-specific ex vivo platforms.

OBJECTIVE: Using STAT1-GoF as an example IEI, we investigated the potential of patient-derived expanded potential stem cells (EPSC) as an ex vivo platform for disease modeling and personalized treatment.

METHODS: We generated EPSC derived from individual STAT1-GoF patients. STAT1 mutations were confirmed with Sanger sequencing. Functional testing including STAT1 phosphorylation/dephosphorylation and gene expression with or without Janus activating kinase inhibitors were performed. Functional tests were repeated on EPSC lines with GoF mutations repaired by CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) editing.

RESULTS: EPSC were successfully reprogrammed from STAT1-GoF patients and expressed the same pluripotent makers as controls, with distinct morphologic differences. Patient-derived EPSC recapitulated the functional abnormalities of index STAT1-GoF patients with STAT1 hyperphosphorylation and increased expression of STAT1 and its downstream genes (IRF1, APOL6, and OAS1) after IFN-γ stimulation. Addition of ruxolitinib and baricitinib inhibited STAT1 hyperactivation in STAT1-GoF EPSC in a dose-dependent manner, which was not observed with tofacitinib. Corrected STAT1 phosphorylation and downstream gene expression were observed among repaired STAT1-GoF EPSC cell lines.

CONCLUSION: This proof-of-concept study demonstrates the potential of our patient-derived EPSC platform to model STAT1-GoF. We propose this platform when researching, recapitulating, and repairing other IEI in the future.

PMID:38072195 | DOI:10.1016/j.jaci.2023.11.914

Employment of a high throughput functional assay to define the critical factors that influence vaccine induced cross-variant neutralizing antibodies for SARS-CoV-2

Sat, 09/12/2023 - 11:00

Sci Rep. 2023 Dec 9;13(1):21810. doi: 10.1038/s41598-023-49231-w.

ABSTRACT

The scale and duration of neutralizing antibody responses targeting SARS-CoV-2 viral variants represents a critically important serological parameter that predicts protective immunity for COVID-19. In this study, we describe the development and employment of a new functional assay that measures neutralizing antibodies for SARS-CoV-2 and present longitudinal data illustrating the impact of age, sex and comorbidities on the kinetics and strength of vaccine-induced antibody responses for key variants in an Asian volunteer cohort. We also present an accurate quantitation of serological responses for SARS-CoV-2 that exploits a unique set of in-house, recombinant human monoclonal antibodies targeting the viral Spike and nucleocapsid proteins and demonstrate a reduction in neutralizing antibody titres across all groups 6 months post-vaccination. We also observe a marked reduction in the serological binding activity and neutralizing responses targeting recently newly emerged Omicron variants including XBB 1.5 and highlight a significant increase in cross-protective neutralizing antibody responses following a third dose (boost) of vaccine. These data illustrate how key virological factors such as immune escape mutations combined with host demographic factors such as age and sex of the vaccinated individual influence the strength and duration of cross-protective serological immunity for COVID-19.

PMID:38071323 | DOI:10.1038/s41598-023-49231-w

Selective digestive tract decontamination to prevent healthcare associated infections in critically ill children: the PICNIC multicentre randomised pilot clinical trial

Fri, 08/12/2023 - 11:00

Sci Rep. 2023 Dec 7;13(1):21668. doi: 10.1038/s41598-023-46232-7.

ABSTRACT

Healthcare-associated infections (HCAIs) are a major cause of morbidity and mortality in critically ill children. Data from adult studies suggest Selective Decontamination of the Digestive tract (SDD) may reduce the incidence of HCAIs and improve survival. There are no data from randomised clinical trials in the paediatric setting. An open label, parallel group pilot cRCT and mixed-methods perspectives study was conducted in six paediatric intensive care units (PICUs) in England. Participants were children (> 37 weeks corrected gestational age, up to 16 years) requiring mechanical ventilation expected to last for at least 48 h. Sites undertook standard care for a period of 9 weeks and were randomised into 3 sites which continued standard care and 3 where SDD was incorporated into infection control practice for eligible children. Interviews and focus groups were conducted for parents and staff working in PICU. 434 children fulfilled eligibility criteria, of whom 368 (85%) were enrolled. This included 207 in the baseline phase (Period One) and 161 in the intervention period (Period Two). In sites delivering SDD, the majority (98%) of children received at least one dose of SDD and of these, 68% commenced within the first 6 h. Whilst admission swabs were collected in 91% of enrolled children, consent for the collection of additional swabs was low (44%). Recruited children were representative of the wider PICU population. Overall, 3.6 children/site/week were recruited compared with the potential recruitment rate for a definitive cRCT of 3 children/site/week, based on data from all UK PICUs. Parents (n = 65) and staff (n = 44) were supportive of the aims of the study, suggesting adaptations for a larger definitive trial including formulation and administration of SDD paste, approaches to consent and ecology monitoring. Stakeholders identified preferred clinical outcomes, focusing on complications of critical illness and quality-of-life. A definitive cRCT in SDD to prevent HCAIs in critically ill children is feasible but should include adaptations to ecology monitoring along with the dosing schedule and packaging into a paediatric specific format. A definitive study is supported by the findings with adaptations to ecology monitoring and SDD administration.Trial Registration: ISRCTN40310490 Registered 30/10/2020.

PMID:38066012 | DOI:10.1038/s41598-023-46232-7

Quantitative proteomics defines mechanisms of antiviral defence and cell death during modified vaccinia Ankara infection

Fri, 08/12/2023 - 11:00

Nat Commun. 2023 Dec 8;14(1):8134. doi: 10.1038/s41467-023-43299-8.

ABSTRACT

Modified vaccinia Ankara (MVA) virus does not replicate in human cells and is the vaccine deployed to curb the current outbreak of mpox. Here, we conduct a multiplexed proteomic analysis to quantify >9000 cellular and ~80% of viral proteins throughout MVA infection of human fibroblasts and macrophages. >690 human proteins are down-regulated >2-fold by MVA, revealing a substantial remodelling of the host proteome. >25% of these MVA targets are not shared with replication-competent vaccinia. Viral intermediate/late gene expression is necessary for MVA antagonism of innate immunity, and suppression of interferon effectors such as ISG20 potentiates virus gene expression. Proteomic changes specific to infection of macrophages indicate modulation of the inflammatory response, including inflammasome activation. Our approach thus provides a global view of the impact of MVA on the human proteome and identifies mechanisms that may underpin its abortive infection. These discoveries will prove vital to design future generations of vaccines.

PMID:38065956 | DOI:10.1038/s41467-023-43299-8

Implementing early rehabilitation and mobilisation for children in UK paediatric intensive care units: the PERMIT feasibility study

Fri, 08/12/2023 - 11:00

Health Technol Assess. 2023 Nov;27(27):1-155. doi: 10.3310/HYRW5688.

ABSTRACT

BACKGROUND: Early rehabilitation and mobilisation encompass patient-tailored interventions, delivered within intensive care, but there are few studies in children and young people within paediatric intensive care units.

OBJECTIVES: To explore how healthcare professionals currently practise early rehabilitation and mobilisation using qualitative and quantitative approaches; co-design the Paediatric Early Rehabilitation and Mobilisation during InTensive care manual of early rehabilitation and mobilisation interventions, with primary and secondary patient-centred outcomes; explore feasibility and acceptability of implementing the Paediatric Early Rehabilitation and Mobilisation during InTensive care manual within three paediatric intensive care units.

DESIGN: Mixed-methods feasibility with five interlinked studies (scoping review, survey, observational study, codesign workshops, feasibility study) in three phases.

SETTING: United Kingdom paediatric intensive care units.

PARTICIPANTS: Children and young people aged 0-16 years remaining within paediatric intensive care on day 3, their parents/guardians and healthcare professionals.

INTERVENTIONS: In Phase 3, unit-wide implementation of manualised early rehabilitation and mobilisation.

MAIN OUTCOME MEASURES: Phase 1 observational study: prevalence of any early rehabilitation and mobilisation on day 3. Phase 3 feasibility study: acceptability of early rehabilitation and mobilisation intervention; adverse events; acceptability of study design; acceptability of outcome measures.

DATA SOURCES: Searched Excerpta Medica Database, Cumulative Index to Nursing and Allied Health Literature, MEDLINE, PEDro, Open grey and Cochrane CENTRAL databases.

REVIEW METHODS: Narrative synthesis.

RESULTS: In the scoping review we identified 36 full-text reports evaluating rehabilitation initiated within 7 days of paediatric intensive care unit admission, outlining non-mobility and mobility early rehabilitation and mobilisation interventions from 24 to 72 hours and delivered twice daily. With the survey, 124/191 (65%) responded from 26/29 (90%) United Kingdom paediatric intensive care units; the majority considered early rehabilitation and mobilisation a priority. The observational study followed 169 patients from 15 units; prevalence of any early rehabilitation and mobilisation on day 3 was 95.3%. We then developed a manualised early rehabilitation and mobilisation intervention informed by current evidence, experience and theory. All three sites implemented the Paediatric Early Rehabilitation and Mobilisation during InTensive care manual successfully, recruited to target (30 patients recruited) and followed up the patients until day 30 or discharge; 21/30 parents consented to complete additional outcome measures.

LIMITATIONS: The findings represent the views of National Health Service staff but may not be generalisable. We were unable to conduct workshops and interviews with children, young people and parents to support the Paediatric Early Rehabilitation and Mobilisation during InTensive care manual development due to pandemic restrictions.

CONCLUSIONS: A randomised controlled trial is recommended to assess the effectiveness of the manualised early rehabilitation and mobilisation intervention.

FUTURE WORK: A definitive cluster randomised trial of early rehabilitation and mobilisation in paediatric intensive care requires selection of outcome measure and health economic evaluation.

STUDY REGISTRATION: The study is registered as PROSPERO CRD42019151050. The Phase 1 observational study is registered Clinicaltrials.gov NCT04110938 (Phase 1) (registered 1 October 2019) and the Phase 3 feasibility study is registered NCT04909762 (Phase 3) (registered 2 June 2021).

FUNDING: This award was funded by the National Institute for Health and Care Research (NIHR) Health Technology Assessment programme (NIHR award ref: 17/21/06) and is published in full in Health Technology Assessment; Vol. 27, No. 27. See the NIHR Funding and Awards website for further award information.

PMID:38063184 | DOI:10.3310/HYRW5688