Benchmarking UMI-aware and standard variant callers for low frequency ctDNA variant detection.

Background: Circulating tumour DNA (ctDNA) is a subset of cell free DNA (cfDNA) released by tumour cells into the bloodstream. Circulating tumour DNA has shown great potential as a biomarker to inform treatment in cancer patients. Collecting ctDNA is minimally invasive and reflects the entire genetic makeup of a patient's cancer.

An Interpretable Classification Model Using Gluten-Specific TCR Sequences Shows Diagnostic Potential in Coeliac Disease.

Coeliac disease (CeD) is a T-cell mediated enteropathy triggered by dietary gluten which remains substantially under-diagnosed around the world. The diagnostic gold-standard requires histological assessment of intestinal biopsies taken at endoscopy while consuming a gluten-containing diet. However, there is a lack of concordance between pathologists in histological assessment, and both endoscopy and gluten challenge are burdensome and unpleasant for patients.

Genomic profiling of idiopathic peri-hilar cholangiocarcinoma reveals new targets and mutational pathways.

Peri-hilar cholangiocarcinoma (pCCA) is chemorefractory and limited genomic analyses have been undertaken in Western idiopathic disease. We undertook comprehensive genomic analyses of a U.K.

Genetic Insights from Consanguineous Cardiomyopathy Families.

Inherited cardiomyopathies are a prevalent cause of heart failure and sudden cardiac death. Both hypertrophic (HCM) and dilated cardiomyopathy (DCM) are genetically heterogeneous and typically present with an autosomal dominant mode of transmission. Whole exome sequencing and autozygosity mapping was carried out in eight un-related probands from consanguineous Middle Eastern families presenting with HCM/DCM followed by bioinformatic and co-segregation analysis to predict the potential pathogenicity of candidate variants.

DECoN: A Detection and Visualization Tool for Exonic Copy Number Variants.

Detection of copy number variants from targeted sequencing, including whole-exome sequencing, can be particularly difficult since the break points of the CNV are not always captured. Here we describe DECoN, a software tool which uses changes in read depth to identify CNVs that affect whole exons. It is optimized for clinical use and allows for interactive visualization of CNVs identified.

Utility of Bulk T-Cell Receptor Repertoire Sequencing Analysis in Understanding Immune Responses to COVID-19.

Measuring immunity to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 19 (COVID-19), can rely on antibodies, reactive T cells and other factors, with T-cell-mediated responses appearing to have greater sensitivity and longevity. Because each T cell carries an essentially unique nucleic acid sequence for its T-cell receptor (TCR), we can interrogate sequence data derived from DNA or RNA to assess aspects of the immune response. This review deals with the utility of bulk, rather than single-cell, sequencing of TCR repertoires, considering the importance of study design, in terms of cohort selection, laboratory methods and analysis.

Predicting the animal hosts of coronaviruses from compositional biases of spike protein and whole genome sequences through machine learning.

The COVID-19 pandemic has demonstrated the serious potential for novel zoonotic coronaviruses to emerge and cause major outbreaks. The immediate animal origin of the causative virus, SARS-CoV-2, remains unknown, a notoriously challenging task for emerging disease investigations. Coevolution with hosts leads to specific evolutionary signatures within viral genomes that can inform likely animal origins.

Use of machine learning to identify a T cell response to SARS-CoV-2.

The identification of SARS-CoV-2-specific T cell receptor (TCR) sequences is critical for understanding T cell responses to SARS-CoV-2. Accordingly, we reanalyze publicly available data from SARS-CoV-2-recovered patients who had low-severity disease (n = 17) and SARS-CoV-2 infection-naive (control) individuals (n = 39). Applying a machine learning approach to TCR beta (TRB) repertoire data, we can classify patient/control samples with a training sensitivity, specificity, and accuracy of 88.

Classification of intestinal T-cell receptor repertoires using machine learning methods can identify patients with coeliac disease regardless of dietary gluten status.

In coeliac disease (CeD), immune-mediated small intestinal damage is precipitated by gluten, leading to variable symptoms and complications, occasionally including aggressive T-cell lymphoma. Diagnosis, based primarily on histopathological examination of duodenal biopsies, is confounded by poor concordance between pathologists and minimal histological abnormality if insufficient gluten is consumed. CeD pathogenesis involves both CD4 T-cell-mediated gluten recognition and CD8 and γδ T-cell-mediated inflammation, with a previous study demonstrating a permanent change in γδ T-cell populations in CeD.

Inferring B cell specificity for vaccines using a Bayesian mixture model.

Background: Vaccines have greatly reduced the burden of infectious disease, ranking in their impact on global health second only after clean water. Most vaccines confer protection by the production of antibodies with binding affinity for the antigen, which is the main effector function of B cells. This results in short term changes in the B cell receptor (BCR) repertoire when an immune response is launched, and long term changes when immunity is conferred.

Use of Transnasal Humidified Rapid-Insufflation Ventilatory Exchange for Emergent Surgical Tracheostomy: A Case Report.

Transnasal humidified rapid-insufflation ventilatory exchange (THRIVE) is a novel airway technique that utilizes high-flow humidified nasal oxygen. It can extend apnea time and maintain oxygen saturation. Here we report the use of THRIVE in a 35-year-old man who required emergent surgical tracheostomy for a clinically relevant compromised airway secondary to acute supraglottic and glottic pathology.

Accurate clinical detection of exon copy number variants in a targeted NGS panel using DECoN.

Targeted next generation sequencing (NGS) panels are increasingly being used in clinical genomics to increase capacity, throughput and affordability of gene testing. Identifying whole exon deletions or duplications (termed exon copy number variants, 'exon CNVs') in exon-targeted NGS panels has proved challenging, particularly for single exon CNVs.  We developed a tool for the Detection of Exon Copy Number variants (DECoN), which is optimised for analysis of exon-targeted NGS panels in the clinical setting.

B-cell repertoire dynamics after sequential hepatitis B vaccination and evidence for cross-reactive B-cell activation.

Background: A diverse B-cell repertoire is essential for recognition and response to infectious and vaccine antigens. High-throughput sequencing of B-cell receptor (BCR) genes can now be used to study the B-cell repertoire at great depth and may shed more light on B-cell responses than conventional immunological methods. Here, we use high-throughput BCR sequencing to provide novel insight into B-cell dynamics following a primary course of hepatitis B vaccination.

Analysis of B Cell Repertoire Dynamics Following Hepatitis B Vaccination in Humans, and Enrichment of Vaccine-specific Antibody Sequences.

Generating a diverse B cell immunoglobulin repertoire is essential for protection against infection. The repertoire in humans can now be comprehensively measured by high-throughput sequencing. Using hepatitis B vaccination as a model, we determined how the total immunoglobulin sequence repertoire changes following antigen exposure in humans, and compared this to sequences from vaccine-specific sorted cells.

The Diversity and Molecular Evolution of B-Cell Receptors during Infection.

B-cell receptors (BCRs) are membrane-bound immunoglobulins that recognize and bind foreign proteins (antigens). BCRs are formed through random somatic changes of germline DNA, creating a vast repertoire of unique sequences that enable individuals to recognize a diverse range of antigens. After encountering antigen for the first time, BCRs undergo a process of affinity maturation, whereby cycles of rapid somatic mutation and selection lead to improved antigen binding.

In-Depth Assessment of Within-Individual and Inter-Individual Variation in the B Cell Receptor Repertoire.

High-throughput sequencing of the B cell receptor (BCR) repertoire can provide rapid characterization of the B cell response in a wide variety of applications in health, after vaccination and in infectious, inflammatory and immune-driven disease, and is starting to yield clinical applications. However, the interpretation of repertoire data is compromised by a lack of studies to assess the intra and inter-individual variation in the BCR repertoire over time in healthy individuals. We applied a standardized isotype-specific BCR repertoire deep sequencing protocol to a single highly sampled participant, and then evaluated the method in 9 further participants to comprehensively describe such variation.

BCR repertoire sequencing: different patterns of B-cell activation after two Meningococcal vaccines.

Next-generation sequencing was used to investigate the B-cell receptor heavy chain transcript repertoire of different B-cell subsets (naive, marginal zone (MZ), immunoglobulin M (IgM) memory and IgG memory) at baseline, and of plasma cells (PCs) 7 days following administration of serogroup ACWY meningococcal polysaccharide and protein-polysaccharide conjugate vaccines. Baseline B-cell subsets could be distinguished from each other using a small number of repertoire properties (clonality, mutation from germline and complementarity-determining region 3 (CDR3) length) that were conserved between individuals. However, analyzing the CDR3 amino-acid sequence (which is particularly important for antigen binding) of the baseline subsets showed few sequences shared between individuals.

Dynamic Bayesian clustering.

Clusters of time series data may change location and memberships over time; in gene expression data, this occurs as groups of genes or samples respond differently to stimuli or experimental conditions at different times. In order to uncover this underlying temporal structure, we consider dynamic clusters with time-dependent parameters which split and merge over time, enabling cluster memberships to change. These interesting time-dependent structures are useful in understanding the development of organisms or complex organs, and could not be identified using traditional clustering methods.

How we do it: training in airway management for a head and neck unit.

Experience and confidence in the management of the airway is highly variable among junior surgical trainees, who are usually the first on scene when problems arise, particularly out of hours. Juniors must possess the skills required to recognise and institute appropriate management in an airway emergency. We describe a local training programme, an airway equipment trolley, and a protocol for recognition, stabilisation, and management, in case of an airway emergency.