Evaluation of the feasibility, diagnostic yield, and clinical utility of rapid genome sequencing in infantile epilepsy (Gene-STEPS): an international, multicentre, pilot cohort study

BACKGROUND: Most neonatal and infantile-onset epilepsies have presumed genetic aetiologies, and early genetic diagnoses have the potential to inform clinical management and improve outcomes. We therefore aimed to determine the feasibility, diagnostic yield, and clinical utility of rapid genome sequencing in this population. METHODS: We conducted an international, multicentre, cohort study (Gene-STEPS), which is a pilot study of the International Precision Child Health Partnership (IPCHiP). IPCHiP is a consortium of four paediatric centres with tertiary-level subspecialty services in Australia, Canada, the UK, and the USA. We recruited infants with new-onset epilepsy or complex febrile seizures from IPCHiP centres, who were younger than 12 months at seizure onset. We excluded infants with simple febrile seizures, acute provoked seizures, known acquired cause, or known genetic cause. Blood samples were collected from probands and available biological parents. Clinical data were collected from medical records, treating clinicians, and parents. Trio genome sequencing was done when both parents were available, and duo or singleton genome sequencing was done when one or neither parent was available. Site-specific protocols were used for DNA extraction and library preparation. Rapid genome sequencing and analysis was done at clinically accredited laboratories, and results were returned to families. We analysed summary statistics for cohort demographic and clinical characteristics and the timing, diagnostic yield, and clinical impact of rapid genome sequencing. FINDINGS: Between Sept 1, 2021, and Aug 31, 2022, we enrolled 100 infants with new-onset epilepsy, of whom 41 (41%) were girls and 59 (59%) were boys. Median age of seizure onset was 128 days (IQR 46-192). For 43 (43% [binomial distribution 95% CI 33-53]) of 100 infants, we identified genetic diagnoses, with a median time from seizure onset to rapid genome sequencing result of 37 days (IQR 25-59). Genetic diagnosis was associated with neonatal seizure onset versus infantile seizure onset (14 [74%] of 19 vs 29 [36%] of 81; p=0·0027), referral setting (12 [71%] of 17 for intensive care, 19 [44%] of 43 non-intensive care inpatient, and 12 [28%] of 40 outpatient; p=0·0178), and epilepsy syndrome (13 [87%] of 15 for self-limited epilepsies, 18 [35%] of 51 for developmental and epileptic encephalopathies, 12 [35%] of 34 for other syndromes; p=0·001). Rapid genome sequencing revealed genetic heterogeneity, with 34 unique genes or genomic regions implicated. Genetic diagnoses had immediate clinical utility, informing treatment (24 [56%] of 43), additional evaluation (28 [65%]), prognosis (37 [86%]), and recurrence risk counselling (all cases). INTERPRETATION: Our findings support the feasibility of implementation of rapid genome sequencing in the clinical care of infants with new-onset epilepsy. Longitudinal follow-up is needed to further assess the role of rapid genetic diagnosis in improving clinical, quality-of-life, and economic outcomes. FUNDING: American Academy of Pediatrics, Boston Children's Hospital Children's Rare Disease Cohorts Initiative, Canadian Institutes of Health Research, Epilepsy Canada, Feiga Bresver Academic Foundation, Great Ormond Street Hospital Charity, Medical Research Council, Murdoch Children's Research Institute, National Institute of Child Health and Human Development, National Institute for Health and Care Research Great Ormond Street Hospital Biomedical Research Centre, One8 Foundation, Ontario Brain Institute, Robinson Family Initiative for Transformational Research, The Royal Children's Hospital Foundation, University of Toronto McLaughlin Centre

Association of Marek's Disease induced immunosuppression with activation of a novel regulatory T cells in chickens.

Marek’s Disease Virus (MDV) is an alphaherpesvirus that infects chickens, transforms CD4+ T cells and causes deadly lymphomas. In addition, MDV induces immunosuppression early during infection by inducing cell death of the infected lymphocytes, and potentially due to activation of regulatory T (Treg)-cells. Furthermore, immunosuppression also occurs during the transformation phase of the disease; however, it is still unknown how the disease can suppress immune response prior or after lymphoma formation. Here, we demonstrated that chicken TGF-beta+ Treg cells are found in different lymphoid tissues, with the highest levels found in the gut-associated lymphoid tissue (cecal tonsil: CT), fostering an immune-privileged microenvironment exerted by TGF-beta. Surprisingly, significantly higher frequencies of TGF-beta+ Treg cells are found in the spleens of MDV-susceptible chicken lines compared to the resistant line, suggesting an association between TGF-beta+ Treg cells and host susceptibility to lymphoma formation. Experimental infection with a virulent MDV elevated the levels of TGF-beta+ Treg cells in the lungs as early as 4 days post infection, and during the transformation phase of the disease in the spleens. In contrast to TGF-beta+ Treg cells, the levels of CD4+CD25+ T cells remained unchanged during the infection and transformation phase of the disease. Furthermore, our results demonstrate that the induction of TGF-beta+ Treg cells is associated with pathogenesis of the disease, as the vaccine strain of MDV did not induce TGF-beta+ Treg cells. Similar to human haematopoietic malignant cells, MDV-induced lymphoma cells expressed high levels of TGF-beta but very low levels of TGF-beta receptor I and II genes. The results confirm that COX-2/ PGE2 pathway is involved in immunosuppression induced by MDV-lymphoma cells. Taken together, our results revealed a novel TGF-beta+ Treg subset in chickens that is activated during MDV infection and tumour formation.Biotechnology and Biological Sciences Research Counci

1,25-Dihydroxyvitamin D3 modulates the phenotype and function of Monocyte derived dendritic cells in cattle

Abstract Background The active form of the vitamin D3, 1,25-Dihydroxyvitamin D3 (1,25-(OH)2D3) has been shown to have major effects not only on physiological processes but also on the regulation of the immune system of vertebrates. Dendritic cells are specialised antigen presenting cells which are in charge of the initiation of T-cell dependant immune responses and as such are key regulators of responses towards pathogens. In this study we set out to evaluate the effects of 1,25-(OH)2D3 on the phenotype of cattle monocyte-derived dendritic cells (MoDCs) and how the conditioning with this vitamin affects the function of these myeloid cells. Results MoDCs were generated from CD14+ monocytes with bovine IL-4 and GM-CSF with or without 1,25-(OH)2D3 supplementation for 10 days. Vitamin D conditioned MoDCs showed a reduced expression of co-stimulatory and antigen presenting molecules, as well as a reduced capability of endocytose ovalbumin. Furthermore, the capacity of MoDCs to induce proliferation in an allogeneic mixed leukocyte reaction was abolished when MoDCs were generated in presence of 1,25-(OH)2D3. LPS induced maturation of 1,25-(OH)2D3conditioned MoDCs resulted in lower secretion of IL-12 and higher IL-10 than that observed in MoDCs. Conclusions The typical immunotolerant phenotype observed in cattle DCs after exposure to 1,25-(OH)2D3 has a significant effect on the functionality of these immune cells, inhibiting the T-cell stimulatory capacity of MoDCs. This could have profound implications on how the bovine immune system deals with pathogens, particularly in diseases such as tuberculosis or paratuberculosis

Development and validation of a search filter to identify equity-focused studies: reducing the number needed to screen

Search strategies and validation set papers. Contains validated search strategies, the number of records retrieved, and the MEDLINE validation set of papers. (PDF 722 kb

Metallic iron nanoparticles intensified production by spinning disk reactor. Optimization and fluid dynamics modelling

The aim of this work was to investigate the optimization of iron nanoparticles production by spinning disk reactor. The influence of the two main operating parameters, i.e. rotational velocity and feed injection point position was investigated through evaluating the particle size distribution, the X-Ray powder diffraction spectra and metallic iron percentage production. Results showed that increasing both rotational velocity and the distance of reagents injection feed points from the disk centre led, to the production of metallic iron nanoparticles characterized by lower mean size. In particular, the optimal rotational velocity was found to be 1400 rpm whereas the optimal distance of injection feed point from disk centre was found to be 3.5 cm. According to these operating parameter values it was possible to obtain monodisperse nanoparticles, characterized by a mean size of 28 ± 2.1 nm and a production in the range 0.24–24 kg/day depending on the initial Fe(II) concentration. The results were then interpreted in light of three fluid dynamics models able to simulate the rotating thin liquid film on the surface of the spinning disk reactor. The applicability of Nusselt model was also investigated and discussed

The safety and effectiveness of a nurse led cardioversion service under sedation

Objective: To assess the safety and effectiveness of nurse led elective cardioversion of atrial fibrillation under sedation. Design: Prospective, longitudinal study. Setting: Cardiac catheterisation laboratory and recovery area of a district general hospital. Patients: 300 patients referred for elective cardioversion of persistent atrial fibrillation. Interventions: Pre-procedure evaluations (history, physical examination, blood tests), consent, sedation administration, cardioversions, and post-procedure monitoring until discharge by advanced life support certified coronary care unit nurses trained in the techniques. A doctor was immediately available if required but not present. Main outcome measures: Success rates at discharge and at six weeks, energy delivered, number of shocks, dose of sedation, immediate, 24, and 48 hour patient perceptions, complications, waiting times, and cost effectiveness. Results: Cardioversion success rate was 87% at discharge and 48% at six weeks. Mean (SD) cumulative energy was 497 (282) J and number of shocks 1.6 (0.8). Mean (SD) dose of sedation was 23 (9) mg intravenous diazepam. No patient required reversal of sedation, airway support, or medical intervention. Ninety eight per cent of patients had no pain or recall of the procedure. Four patients who were adequately anticoagulated experienced embolic phenomena. Ninety eight per cent of patients would repeat the procedure if necessary. Without requirement for a physician or anaesthetist, waiting times for elective cardioversion fell from three months to under four weeks. There was a significant reduction in the estimated cost of the procedure from £337 with general anaesthesia to £130 with nurse led sedation and cardioversion (p < 0.001). Conclusion: With appropriate training, a nurse led cardioversion service with sedation is safe, effective, well tolerated, and cost efficient

The relationship between high-frequency right ventricular pacing and paroxysmal atrial fibrillation burden

Right ventricular pacing increases the risk of persistent atrial fibrillation (AF) in the long term. The effects of right ventricular pacing on paroxysmal AF (PAF) are unknown. The aim was to examine the effect of right ventricular pacing on AF burden (AFB) in patients with symptomatic drug-resistant PAF. Pooled analysis of pacemaker-derived counters and AF diagnostic data from the Atrial Fibrillation Therapy (AFT) and Pacemaker Atrial Fibrillation Suppression (PAFS) randomized anti-AF pacemaker algorithm trials were used.Five hundred and fifty-four patients from the AFT (n = 372) and PAFS (n = 182) were studied. The individual percentages of pacing, Atrial Sense Ventricular Pace (ASVP), Atrial Pace Ventricular Pace (APVP), and Atrial Pace Ventricular Sense (APVS) as well as total ventricular pacing during synchronous rhythm (VPinSR, %) were examined for an effect on AFB. Three hundred and twenty-one (AFT, age 64 +/- 11, 55% male) and 79 (PAFS, age 71 +/- 8, 54% male) patients had complete data for analysis. Increased VPinSR was weakly associated with an increased AFB (effect size-10% VPinSR increased AFB by only 0.03%) in AFT (P = 0.04) but not PAFS (P = 0.98) or the pooled analysis (P = 0.95). None of the synchronous paced modalities (ASVP, APVP, APVS) significantly increased AFB compared with sinus rhythm (Atrial Sense Ventricular Sense) (P = ns).No pacing modality, atrial or ventricular, had a significant effect on AFB. On the basis of these data, the detrimental effect of high-frequency right ventricular pacing on AFB in paced PAF patients, unlike with persistent AF, appears to be minimal in the short term