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

Comparative mitochondrial proteomics: perspective in human diseases

Mitochondria are the most complex and the most important organelles of eukaryotic cells, which are involved in many cellular processes, including energy metabolism, apoptosis, and aging. And mitochondria have been identified as the "hot spot" by researchers for exploring relevant associated dysfunctions in many fields. The emergence of comparative proteomics enables us to have a close look at the mitochondrial proteome in a comprehensive and effective manner under various conditions and cellular circumstances. Two-dimensional electrophoresis combined with mass spectrometry is still the most popular techniques to study comparative mitochondrial proteomics. Furthermore, many new techniques, such as ICAT, MudPIT, and SILAC, equip researchers with more flexibilities inselecting proper methods. This article also reviews the recent development of comparative mitochondrial proteomics on diverse human diseases. And the results of mitochondrial proteomics enhance a better understanding of the pathogenesis associated with mitochondria and provide promising therapeutic targets

Is biotechnology (more) acceptable when it enables a reduction in phytosanitary treatments? A European comparison of the acceptability of transgenesis and cisgenesis

Reduced pesticide use is one of the reasons given by Europeans for accepting new genetic engineering techniques. According to the advocates of these techniques, consumers are likely to embrace the application of cisgenesis to apple trees. In order to verify the acceptability of these techniques, we estimate a Bayesian multilevel structural equation model, which takes into account the multidimensional nature of acceptability and individual, national, and European effects, using data from the Eurobarometer 2010 73.1 on science. The results underline the persistence of clear differences between European countries and whilst showing considerable defiance, a relatively wider acceptability of vertical gene transfer as a means of reducing phytosanitary treatments, compared to horizontal transfer

Crowdsourcing Coordination: A Review and Research Agenda for Crowdsourcing Coordination Used for Macro-tasks

Crowdsourcing has become a widely accepted approach to leveraging the skills and expertise of others to accomplish work. Despite the potential of crowdsourcing to tackle complex problems, it has often been used to address simple micro-tasks. To tackle more complex macro-tasks, more attention is needed to better comprehend crowd coordination. Crowd coordination is defined as the synchronization of crowd workers in an attempt to direct and align their efforts in pursuit of a shared goal. The goal of this chapter is to advance our understanding of crowd coordination to tackle complex macro-tasks. To accomplish this, we have three objectives. First, we review popular theories of coordination. Second, we examine the current approaches to crowd coordination in the HCI and CSCW literature. Finally, the chapter identifies shortcomings in the literature and proposes a research agenda directed at advancing our understanding of crowd coordination needed to address complex macro-tasks.National Science Foundation grant CHS-1617820Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/150620/1/Kim and Robert 2019 Preprint Chapter 2.pdfDescription of Kim and Robert 2019 Preprint Chapter 2.pdf : Preprint Versio

Interneuron- and GABAA receptor-specific inhibitory synaptic plasticity in cerebellar purkinje cells

Inhibitory synaptic plasticity is important for shaping both neuronal excitability and network activity. Here we investigate the input and GABA(A) receptor subunit specificity of inhibitory synaptic plasticity by studying cerebellar interneuron-Purkinje cell (PC) synapses. Depolarizing PCs initiated a long-lasting increase in GABA-mediated synaptic currents. By stimulating individual interneurons, this plasticity was observed at somatodendritic basket cell synapses, but not at distal dendritic stellate cell synapses. Basket cell synapses predominantly express β2-subunit-containing GABA(A) receptors; deletion of the β2-subunit ablates this plasticity, demonstrating its reliance on GABA(A) receptor subunit composition. The increase in synaptic currents is dependent upon an increase in newly synthesized cell surface synaptic GABA(A) receptors and is abolished by preventing CaMKII phosphorylation of GABA(A) receptors. Our results reveal a novel GABA(A) receptor subunit- and input-specific form of inhibitory synaptic plasticity that regulates the temporal firing pattern of the principal output cells of the cerebellum

A922 Sequential measurement of 1 hour creatinine clearance (1-CRCL) in critically ill patients at risk of acute kidney injury (AKI)

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Post-transcriptional regulation of virulence gene expression in enterohaemorrhagic E. coli

Enterohaemorrhagic E. coli (EHEC) is a significant foodborne pathogen responsible for outbreaks of haemorrhagic colitisand haemolytic uremic syndrome (HUS). Renal tissue destruction caused by HUS is caused by the release of Shiga toxins (Stx) encoded on lambdoid bacteriophage (StxΦ). Toxin release is dependent on bacteriophage lytic induction and can be triggered by antibiotics. New treatments for EHEC infections are required and this thesis examines the contribution of posttranscriptional regulation of Stx and virulence genes mediated by regulatory small RNAs.The EHEC haem receptor, chuA, is regulated in response to iron availability and temperature. Here, chuA was found tobe subject to Rho-termination and directly activated by the Crp-cAMP regulated sRNA, CyaR, in a temperatureindependent manner. This reveals two additional layers of regulation employed by EHEC to ensure this haem receptor isonly expressed within a host and suggests that chuA employs a transcriptional and post-transcriptional AND/OR-logic gate to integrate multiple signals that indicate host infection.Transcriptome-wide Hfq-binding sites have shown that the StxΦ transcribed at least 11 regulatory small RNAs. Here, anovel small RNA termed StxS was found to be transcribed from the late phage and Shiga toxin promoter PR’ duringlysogeny. This transcript is processed by the endoribonuclease RNase E into a functional sRNA that activates the stresssigma factor RpoS and represses Stx production. These demonstrate how the StxΦ modulates host stress responses and fitness through post-transcriptional regulation. The PR’-tR’ organization critical for transcription of StxS is conserved in lambdoid bacteriophage, suggesting that PR’ may be the origin of an assortment of sRNAs that regulate bacterialpathogenicity and fitness. RNA-binding proteins (RBPs) play a key role in post-transcriptional regulation. Total RNA associated protein purification(TRAPP) was used to recover the RNA-binding proteome of EHEC. TRAPP recovered 443 proteins, 35 being EHEC-specific, including the effector proteins EspY2 and EspN. Four of these were confirmed to be RNA-binding through 32P radiolabelling of RNA-protein complexes. This showed that pathogen specific RBPs may play a key role in EHECpathogenesis.Together, these results highlight the complexity of post-transcriptional regulation in EHEC, and how it affects EHECpathogenicity and fitness