Characterizing the Major Structural Variant Alleles of the Human Genome.

In order to provide a comprehensive resource for human structural variants (SVs), we generated long-read sequence data and analyzed SVs for fifteen human genomes. We sequence resolved 99,604 insertions, deletions, and inversions including 2,238 (1.6 Mbp) that are shared among all discovery genomes with an additional 13,053 (6.9 Mbp) present in the majority, indicating minor alleles or errors in the reference. Genotyping in 440 additional genomes confirms the most common SVs in unique euchromatin are now sequence resolved. We report a ninefold SV bias toward the last 5 Mbp of human chromosomes with nearly 55% of all VNTRs (variable number of tandem repeats) mapping to this portion of the genome. We identify SVs affecting coding and noncoding regulatory loci improving annotation and interpretation of functional variation. These data provide the framework to construct a canonical human reference and a resource for developing advanced representations capable of capturing allelic diversity.

The origin and evolution of mutations in acute myeloid leukemia.

Most mutations in cancer genomes are thought to be acquired after the initiating event, which may cause genomic instability and drive clonal evolution. However, for acute myeloid leukemia (AML), normal karyotypes are common, and genomic instability is unusual. To better understand clonal evolution in AML, we sequenced the genomes of M3-AML samples with a known initiating event (PML-RARA) versus the genomes of normal karyotype M1-AML samples and the exomes of hematopoietic stem/progenitor cells (HSPCs) from healthy people. Collectively, the data suggest that most of the mutations found in AML genomes are actually random events that occurred in HSPCs before they acquired the initiating mutation; the mutational history of that cell is "captured" as the clone expands. In many cases, only one or two additional, cooperating mutations are needed to generate the malignant founding clone. Cells from the founding clone can acquire additional cooperating mutations, yielding subclones that can contribute to disease progression and/or relapse.

The Benefits and Risks of Receiving Investigational Solid Tumor Drugs in Randomized Trials : A Systematic Review and Meta-analysis.

BACKGROUND: Many patients participate in cancer trials to access new therapies. The extent to which new treatments produce clinical benefit for trial participants is unclear. PURPOSE: To estimate the progression-free survival (PFS) and overall survival (OS) advantage of assignment to experimental groups in randomized trials for 6 solid tumors. DATA SOURCES: ClinicalTrials.gov was searched for trials of investigational drugs with results posted between 2017 and 2021. STUDY SELECTION: Investigational drugs were defined as those not yet having full approval from the U.S. Food and Drug Administration for the study indication. Trials were included if they were randomized and tested drugs or biologics. DATA EXTRACTION: Data extraction was completed by 2 independent reviewers. Data were pooled using a random-effects model. DATA SYNTHESIS: The sample included 128 trials comprising 141 comparisons of a new drug and a comparator. These comparisons included 47 050 patients. The pooled hazard ratio for PFS was 0.80 (95% CI, 0.75 to 0.85), indicating statistically significant benefit for patients in experimental groups. This corresponded to a median PFS advantage of 1.25 months (CI, 0.80 to 1.68 months). The pooled hazard ratio for OS was 0.92 (CI, 0.88 to 0.95), corresponding to a survival gain of 1.18 months (CI, 0.72 to 1.71 months). The absolute risk for a serious adverse event for comparator group patients was 29.56% (CI, 26.64% to 32.65%), with an increase in risk of 7.40% (CI, 5.66% to 9.14%) for patients in experimental groups. LIMITATIONS: Trials in this sample were heterogeneous. Comparator group interventions were assumed to reflect standard of care. CONCLUSION: Assignment to experimental groups produces statistically significant survival gains. However, the absolute survival gain is small, and toxicity is statistically significantly greater. The findings of this review provide reassuring evidence that patients are not meaningfully disadvantaged by assignment to comparator groups. PRIMARY FUNDING SOURCE: Canadian Institutes of Health Research.

Identification of difficult laryngoscopy using an optimized hybrid architecture.

BACKGROUND: Identification of difficult laryngoscopy is a frequent demand in cervical spondylosis clinical surgery. This work aims to develop a hybrid architecture for identifying difficult laryngoscopy based on new indexes. METHODS: Initially, two new indexes for identifying difficult laryngoscopy are proposed, and their efficacy for predicting difficult laryngoscopy is compared to that of two conventional indexes. Second, a hybrid adaptive architecture with convolutional layers, spatial extraction, and a vision transformer is proposed for predicting difficult laryngoscopy. The proposed adaptive hybrid architecture is then optimized by determining the optimal location for extracting spatial information. RESULTS: The test accuracy of four indexes using simple model is 0.8320. The test accuracy of optimized hybrid architecture using four indexes is 0.8482. CONCLUSION: The newly proposed two indexes, the angle between the lower margins of the second and sixth cervical spines and the vertical direction, are validated to be effective for recognizing difficult laryngoscopy. In addition, the optimized hybrid architecture employing four indexes demonstrates improved efficacy in detecting difficult laryngoscopy. TRIAL REGISTRATION: Ethics permission for this research was obtained from the Medical Scientific Research Ethics Committee of Peking University Third Hospital (IRB00006761-2015021) on 30 March 2015. A well-informed agreement has been received from all participants. Patients were enrolled in this research at the Chinese Clinical Trial Registry ( http://www.chictr.org.cn , identifier: ChiCTR-ROC-16008598) on 6 June 2016.

Evaluating Model Specification When Using the Parametric G-Formula in the Presence of Censoring.

The noniterative conditional expectation (NICE) parametric g-formula can be used to estimate the causal effect of sustained treatment strategies. In addition to identifiability conditions, the validity of the NICE parametric g-formula generally requires the correct specification of models for time-varying outcomes, treatments, and confounders at each follow-up time point. An informal approach for evaluating model specification is to compare the observed distributions of the outcome, treatments, and confounders with their parametric g-formula estimates under the "natural course." In the presence of loss to follow-up, however, the observed and natural-course risks can differ even if the identifiability conditions of the parametric g-formula hold and there is no model misspecification. Here, we describe 2 approaches for evaluating model specification when using the parametric g-formula in the presence of censoring: 1) comparing factual risks estimated by the g-formula with nonparametric Kaplan-Meier estimates and 2) comparing natural-course risks estimated by inverse probability weighting with those estimated by the g-formula. We also describe how to correctly compute natural-course estimates of time-varying covariate means when using a computationally efficient g-formula algorithm. We evaluate the proposed methods via simulation and implement them to estimate the effects of dietary interventions in 2 cohort studies.

A novel sialic acid-binding adhesin present in multiple species contributes to the pathogenesis of Infective endocarditis.

Bacterial binding to platelets is a key step in the development of infective endocarditis (IE). Sialic acid, a common terminal carbohydrate on host glycans, is the major receptor for streptococci on platelets. So far, all defined interactions between streptococci and sialic acid on platelets are mediated by serine-rich repeat proteins (SRRPs). However, we identified Streptococcus oralis subsp. oralis IE-isolates that bind sialic acid but lack SRRPs. In addition to binding sialic acid, some SRRP- isolates also bind the cryptic receptor ß-1,4-linked galactose through a yet unknown mechanism. Using comparative genomics, we identified a novel sialic acid-binding adhesin, here named AsaA (associated with sialic acid adhesion A), present in IE-isolates lacking SRRPs. We demonstrated that S. oralis subsp. oralis AsaA is required for binding to platelets in a sialic acid-dependent manner. AsaA comprises a non-repeat region (NRR), consisting of a FIVAR/CBM and two Siglec-like and Unique domains, followed by 31 DUF1542 domains. When recombinantly expressed, Siglec-like and Unique domains competitively inhibited binding of S. oralis subsp. oralis and directly interacted with sialic acid on platelets. We further demonstrated that AsaA impacts the pathogenesis of S. oralis subsp. oralis in a rabbit model of IE. Additionally, we found AsaA orthologues in other IE-causing species and demonstrated that the NRR of AsaA from Gemella haemolysans blocked binding of S. oralis subsp. oralis, suggesting that AsaA contributes to the pathogenesis of multiple IE-causing species. Finally, our findings provide evidence that sialic acid is a key factor for bacterial-platelets interactions in a broader range of species than previously appreciated, highlighting its potential as a therapeutic target.

A sequence motif within chromatin entry sites directs MSL establishment on the Drosophila X chromosome.

The Drosophila MSL complex associates with active genes specifically on the male X chromosome to acetylate histone H4 at lysine 16 and increase expression approximately 2-fold. To date, no DNA sequence has been discovered to explain the specificity of MSL binding. We hypothesized that sequence-specific targeting occurs at "chromatin entry sites," but the majority of sites are sequence independent. Here we characterize 150 potential entry sites by ChIP-chip and ChIP-seq and discover a GA-rich MSL recognition element (MRE). The motif is only slightly enriched on the X chromosome ( approximately 2-fold), but this is doubled when considering its preferential location within or 3' to active genes (>4-fold enrichment). When inserted on an autosome, a newly identified site can direct local MSL spreading to flanking active genes. These results provide strong evidence for both sequence-dependent and -independent steps in MSL targeting of dosage compensation to the male X chromosome.

Long-read whole genome sequencing reveals HOXD13 alterations in synpolydactyly.

Synpolydactyly 1, also called syndactyly type II (SDTY2), is a genetic limb malformation characterized by polydactyly with syndactyly involving the webbing of the third and fourth fingers, and the fourth and fifth toes. It is caused by heterozygous alterations in HOXD13 with incomplete penetrance and phenotypic variability. In our study, a five-generation family with an SPD phenotype was enrolled in our Rare Disease Genomics Protocol. A comprehensive examination of three generations using Illumina short-read whole-genome sequencing (WGS) did not identify any causative variants. Subsequent WGS using Pacific Biosciences (PacBio) long-read HiFi Circular Consensus Sequencing (CCS) revealed a heterozygous 27-bp duplication in the polyalanine tract of HOXD13. Sanger sequencing of all available family members confirmed that the variant segregates with affected individuals. Reanalysis of an unrelated family with a similar SPD phenotype uncovered a 21-bp (7-alanine) duplication in the same region of HOXD13. Although ExpansionHunter identified these events in most individuals in a retrospective analysis, low sequence coverage due to high GC content in the HOXD13 polyalanine tract makes detection of these events challenging. Our findings highlight the value of long-read WGS in elucidating the molecular etiology of congenital limb malformation disorders.

Accuracy of rapid point-of-care antigen-based diagnostics for SARS-CoV-2: An updated systematic review and meta-analysis with meta-regression analyzing influencing factors.

BACKGROUND: Comprehensive information about the accuracy of antigen rapid diagnostic tests (Ag-RDTs) for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is essential to guide public health decision makers in choosing the best tests and testing policies. In August 2021, we published a systematic review and meta-analysis about the accuracy of Ag-RDTs. We now update this work and analyze the factors influencing test sensitivity in further detail. METHODS AND FINDINGS: We registered the review on PROSPERO (registration number: CRD42020225140). We systematically searched preprint and peer-reviewed databases for publications evaluating the accuracy of Ag-RDTs for SARS-CoV-2 until August 31, 2021. Descriptive analyses of all studies were performed, and when more than 4 studies were available, a random-effects meta-analysis was used to estimate pooled sensitivity and specificity with reverse transcription polymerase chain reaction (RT-PCR) testing as a reference. To evaluate factors influencing test sensitivity, we performed 3 different analyses using multivariable mixed-effects meta-regression models. We included 194 studies with 221,878 Ag-RDTs performed. Overall, the pooled estimates of Ag-RDT sensitivity and specificity were 72.0% (95% confidence interval [CI] 69.8 to 74.2) and 98.9% (95% CI 98.6 to 99.1). When manufacturer instructions were followed, sensitivity increased to 76.3% (95% CI 73.7 to 78.7). Sensitivity was markedly better on samples with lower RT-PCR cycle threshold (Ct) values (97.9% [95% CI 96.9 to 98.9] and 90.6% [95% CI 88.3 to 93.0] for Ct-values <20 and <25, compared to 54.4% [95% CI 47.3 to 61.5] and 18.7% [95% CI 13.9 to 23.4] for Ct-values ≥25 and ≥30) and was estimated to increase by 2.9 percentage points (95% CI 1.7 to 4.0) for every unit decrease in mean Ct-value when adjusting for testing procedure and patients' symptom status. Concordantly, we found the mean Ct-value to be lower for true positive (22.2 [95% CI 21.5 to 22.8]) compared to false negative (30.4 [95% CI 29.7 to 31.1]) results. Testing in the first week from symptom onset resulted in substantially higher sensitivity (81.9% [95% CI 77.7 to 85.5]) compared to testing after 1 week (51.8%, 95% CI 41.5 to 61.9). Similarly, sensitivity was higher in symptomatic (76.2% [95% CI 73.3 to 78.9]) compared to asymptomatic (56.8% [95% CI 50.9 to 62.4]) persons. However, both effects were mainly driven by the Ct-value of the sample. With regards to sample type, highest sensitivity was found for nasopharyngeal (NP) and combined NP/oropharyngeal samples (70.8% [95% CI 68.3 to 73.2]), as well as in anterior nasal/mid-turbinate samples (77.3% [95% CI 73.0 to 81.0]). Our analysis was limited by the included studies' heterogeneity in viral load assessment and sample origination. CONCLUSIONS: Ag-RDTs detect most of the individuals infected with SARS-CoV-2, and almost all (>90%) when high viral loads are present. With viral load, as estimated by Ct-value, being the most influential factor on their sensitivity, they are especially useful to detect persons with high viral load who are most likely to transmit the virus. To further quantify the effects of other factors influencing test sensitivity, standardization of clinical accuracy studies and access to patient level Ct-values and duration of symptoms are needed.

Biomarkers That Correlate with Active Pulmonary Tuberculosis Treatment Response: a Systematic Review and Meta-analysis.

Current WHO recommendations for monitoring treatment response in adult pulmonary tuberculosis (TB) are sputum smear microscopy and/or culture conversion at the end of the intensive phase of treatment. These methods either have suboptimal accuracy or a long turnaround time. There is a need to identify alternative biomarkers to monitor TB treatment response. We conducted a systematic review of active pulmonary TB treatment monitoring biomarkers. We screened 9,739 articles published between 1 January 2008 and 31 December 2020, of which 77 met the inclusion criteria. When studies quantitatively reported biomarker levels, we meta-analyzed the average fold change in biomarkers from pretreatment to week 8 of treatment. We also performed a meta-analysis pooling the fold change since the previous time point collected. A total of 81 biomarkers were identified from 77 studies. Overall, these studies exhibited extensive heterogeneity with regard to TB treatment monitoring study design and data reporting. Among the biomarkers identified, C-reactive protein (CRP), interleukin-6 (IL-6), interferon gamma-induced protein 10 (IP-10), and tumor necrosis factor alpha (TNF-α) had sufficient data to analyze fold changes. All four biomarker levels decreased during the first 8 weeks of treatment relative to baseline and relative to previous time points collected. Based on limited data available, CRP, IL-6, IP-10, and TNF-α have been identified as biomarkers that should be further explored in the context of TB treatment monitoring. The extensive heterogeneity in TB treatment monitoring study design and reporting is a major barrier to evaluating the performance of novel biomarkers and tools for this use case. Guidance for designing and reporting treatment monitoring studies is urgently needed.

Revisiting the g-null Paradox.

The (noniterative conditional expectation) parametric g-formula is an approach to estimating causal effects of sustained treatment strategies from observational data. An often-cited limitation of the parametric g-formula is the g-null paradox: a phenomenon in which model misspecification in the parametric g-formula is guaranteed in some settings consistent with the conditions that motivate its use (i.e., when identifiability conditions hold and measured time-varying confounders are affected by past treatment). Many users of the parametric g-formula acknowledge the g-null paradox as a limitation when reporting results but still require clarity on its meaning and implications. Here, we revisit the g-null paradox to clarify its role in causal inference studies. In doing so, we present analytic examples and a simulation-based illustration of the bias of parametric g-formula estimates under the conditions associated with this paradox. Our results highlight the importance of avoiding overly parsimonious models for the components of the g-formula when using this method.

Detection of brain somatic variation in epilepsy-associated developmental lesions.

OBJECTIVE: Epilepsy-associated developmental lesions, including malformations of cortical development and low-grade developmental tumors, represent a major cause of drug-resistant seizures requiring surgical intervention in children. Brain-restricted somatic mosaicism has been implicated in the genetic etiology of these lesions; however, many contributory genes remain unidentified. METHODS: We enrolled 50 children who were undergoing epilepsy surgery into a translational research study. Resected tissue was divided for clinical neuropathologic evaluation and genomic analysis. We performed exome and RNA sequencing to identify somatic variation and we confirmed our findings using high-depth targeted DNA sequencing. RESULTS: We uncovered candidate disease-causing somatic variation affecting 28 patients (56%), as well as candidate germline variants affecting 4 patients (8%). In agreement with previous studies, we identified somatic variation affecting solute carrier family 35 member A2 (SLC35A2) and mechanistic target of rapamycin kinase (MTOR) pathway genes in patients with focal cortical dysplasia. Somatic gains of chromosome 1q were detected in 30% (3 of 10) of patients with Type I focal cortical dysplasia (FCD)s. Somatic variation in mitogen-activated protein kinase (MAPK) pathway genes (i.e., fibroblast growth factor receptor 1 [FGFR1], FGFR2, B-raf proto-oncogene, serine/threonine kinase [BRAF], and KRAS proto-oncogene, GTPase [KRAS]) was associated with low-grade epilepsy-associated developmental tumors. RNA sequencing enabled the detection of somatic structural variation that would have otherwise been missed, and which accounted for more than one-half of epilepsy-associated tumor diagnoses. Sampling across multiple anatomic regions revealed that somatic variant allele fractions vary widely within epileptogenic tissue. Finally, we identified putative disease-causing variants in genes not yet associated with focal cortical dysplasia. SIGNIFICANCE: These results further elucidate the genetic basis of structural brain abnormalities leading to focal epilepsy in children and point to new candidate disease genes.

Impact of molecular diagnostic tests on diagnostic and treatment delays in tuberculosis: a systematic review and meta-analysis.

BACKGROUND: Countries with high TB burden have expanded access to molecular diagnostic tests. However, their impact on reducing delays in TB diagnosis and treatment has not been assessed. Our primary aim was to summarize the quantitative evidence on the impact of nucleic acid amplification tests (NAAT) on diagnostic and treatment delays compared to that of the standard of care for drug-sensitive and drug-resistant tuberculosis (DS-TB and DR-TB). METHODS: We searched MEDLINE, EMBASE, Web of Science, and the Global Health databases (from their inception to October 12, 2020) and extracted time delay data for each test. We then analysed the diagnostic and treatment initiation delay separately for DS-TB and DR-TB by comparing smear vs Xpert for DS-TB and culture drug sensitivity testing (DST) vs line probe assay (LPA) for DR-TB. We conducted random effects meta-analyses of differences of the medians to quantify the difference in diagnostic and treatment initiation delay, and we investigated heterogeneity in effect estimates based on the period the test was used in, empiric treatment rate, HIV prevalence, healthcare level, and study design. We also evaluated methodological differences in assessing time delays. RESULTS: A total of 45 studies were included in this review (DS = 26; DR = 20). We found considerable heterogeneity in the definition and reporting of time delays across the studies. For DS-TB, the use of Xpert reduced diagnostic delay by 1.79 days (95% CI - 0.27 to 3.85) and treatment initiation delay by 2.55 days (95% CI 0.54-4.56) in comparison to sputum microscopy. For DR-TB, use of LPAs reduced diagnostic delay by 40.09 days (95% CI 26.82-53.37) and treatment initiation delay by 45.32 days (95% CI 30.27-60.37) in comparison to any culture DST methods. CONCLUSIONS: Our findings indicate that the use of World Health Organization recommended diagnostics for TB reduced delays in diagnosing and initiating TB treatment. Future studies evaluating performance and impact of diagnostics should consider reporting time delay estimates based on the standardized reporting framework.

PTEN somatic mutations contribute to spectrum of cerebral overgrowth.

Phosphatase and tensin homologue (PTEN) regulates cell growth and survival through inhibition of the mammalian target of rapamycin (MTOR) signalling pathway. Germline genetic variation of PTEN is associated with autism, macrocephaly and PTEN hamartoma tumour syndromes. The effect of developmental PTEN somatic mutations on nervous system phenotypes is not well understood, although brain somatic mosaicism of MTOR pathway genes is an emerging cause of cortical dysplasia and epilepsy in the paediatric population. Here we report two somatic variants of PTEN affecting a single patient presenting with intractable epilepsy and hemimegalencephaly that varied in clinical severity throughout the left cerebral hemisphere. High-throughput sequencing analysis of affected brain tissue identified two somatic variants in PTEN. The first variant was present in multiple cell lineages throughout the entire hemisphere and associated with mild cerebral overgrowth. The second variant was restricted to posterior brain regions and affected the opposite PTEN allele, resulting in a segmental region of more severe malformation, and the only neurons in which it was found by single-nuclei RNA-sequencing had a unique disease-related expression profile. This study reveals brain mosaicism of PTEN as a disease mechanism of hemimegalencephaly and furthermore demonstrates the varying effects of single- or bi-allelic disruption of PTEN on cortical phenotypes.

Novel morphologic findings in PLAG1-rearranged soft tissue tumors.

Oncogenesis in PLAG1-rearranged tumors often results from PLAG1 transcription factor overexpression driven by promoter-swapping between constitutively expressed fusion partners. PLAG1-rearranged tumors demonstrate diverse morphologies. This study adds to this morphologic heterogeneity by introducing two tumors with PLAG1 rearrangements that display distinct histologic features. The first arose in the inguinal region of a 3-year-old, appeared well-circumscribed with a multinodular pattern, and harbored two fusions: ZFHX4-PLAG1 and CHCHD7-PLAG1. The second arose in the pelvic cavity of a 15-year-old girl, was extensively infiltrative and vascularized with an adipocytic component, and demonstrated a COL3A1-PLAG1 fusion. Both showed low-grade cytomorphology, scarce mitoses, no necrosis, and expression of CD34 and desmin. The ZFHX4-/CHCHD7-PLAG1-rearranged tumor showed no evidence of recurrence after 5 months. By contrast, the COL3A1-PLAG1-rearranged tumor quickly recurred following primary excision with positive margins; subsequent re-excision with adjuvant chemotherapy resulted in no evidence of recurrence after 2 years. While both tumors show overlap with benign and malignant fibroblastic and fibrovascular neoplasms, they also display divergent features. These cases highlight the importance of appropriate characterization in soft tissue tumors with unusual clinical and histologic characteristics.

Gastroblastoma with a novel EWSR1-CTBP1 fusion presenting in adolescence.

Gastroblastomas are rare tumors with a biphasic epithelioid/spindle cell morphology that typically present in early adulthood and have recurrent MALAT1-GLI1 fusions. We describe an adolescent patient with Wiskott-Aldrich syndrome who presented with a large submucosal gastric tumor with biphasic morphology. Despite histologic features consistent with gastroblastoma, a MALAT1-GLI1 fusion was not found in this patient's tumor; instead, comprehensive molecular profiling identified a novel EWSR1-CTBP1 fusion and no other significant genetic alterations. The tumor also overexpressed NOTCH and FGFR by RNA profiling. The novel fusion and expression profile suggest a role for epithelial-mesenchymal transition in this tumor, with potential implications for the pathogenesis of biphasic gastric tumors such as gastroblastoma.

Discovery of clinically relevant fusions in pediatric cancer.

BACKGROUND: Pediatric cancers typically have a distinct genomic landscape when compared to adult cancers and frequently carry somatic gene fusion events that alter gene expression and drive tumorigenesis. Sensitive and specific detection of gene fusions through the analysis of next-generation-based RNA sequencing (RNA-Seq) data is computationally challenging and may be confounded by low tumor cellularity or underlying genomic complexity. Furthermore, numerous computational tools are available to identify fusions from supporting RNA-Seq reads, yet each algorithm demonstrates unique variability in sensitivity and precision, and no clearly superior approach currently exists. To overcome these challenges, we have developed an ensemble fusion calling approach to increase the accuracy of identifying fusions. RESULTS: Our Ensemble Fusion (EnFusion) approach utilizes seven fusion calling algorithms: Arriba, CICERO, FusionMap, FusionCatcher, JAFFA, MapSplice, and STAR-Fusion, which are packaged as a fully automated pipeline using Docker and Amazon Web Services (AWS) serverless technology. This method uses paired end RNA-Seq sequence reads as input, and the output from each algorithm is examined to identify fusions detected by a consensus of at least three algorithms. These consensus fusion results are filtered by comparison to an internal database to remove likely artifactual fusions occurring at high frequencies in our internal cohort, while a "known fusion list" prevents failure to report known pathogenic events. We have employed the EnFusion pipeline on RNA-Seq data from 229 patients with pediatric cancer or blood disorders studied under an IRB-approved protocol. The samples consist of 138 central nervous system tumors, 73 solid tumors, and 18 hematologic malignancies or disorders. The combination of an ensemble fusion-calling pipeline and a knowledge-based filtering strategy identified 67 clinically relevant fusions among our cohort (diagnostic yield of 29.3%), including RBPMS-MET, BCAN-NTRK1, and TRIM22-BRAF fusions. Following clinical confirmation and reporting in the patient's medical record, both known and novel fusions provided medically meaningful information. CONCLUSIONS: The EnFusion pipeline offers a streamlined approach to discover fusions in cancer, at higher levels of sensitivity and accuracy than single algorithm methods. Furthermore, this method accurately identifies driver fusions in pediatric cancer, providing clinical impact by contributing evidence to diagnosis and, when appropriate, indicating targeted therapies.

Evaluation of GRCh38 and de novo haploid genome assemblies demonstrates the enduring quality of the reference assembly.

The human reference genome assembly plays a central role in nearly all aspects of today's basic and clinical research. GRCh38 is the first coordinate-changing assembly update since 2009; it reflects the resolution of roughly 1000 issues and encompasses modifications ranging from thousands of single base changes to megabase-scale path reorganizations, gap closures, and localization of previously orphaned sequences. We developed a new approach to sequence generation for targeted base updates and used data from new genome mapping technologies and single haplotype resources to identify and resolve larger assembly issues. For the first time, the reference assembly contains sequence-based representations for the centromeres. We also expanded the number of alternate loci to create a reference that provides a more robust representation of human population variation. We demonstrate that the updates render the reference an improved annotation substrate, alter read alignments in unchanged regions, and impact variant interpretation at clinically relevant loci. We additionally evaluated a collection of new de novo long-read haploid assemblies and conclude that although the new assemblies compare favorably to the reference with respect to continuity, error rate, and gene completeness, the reference still provides the best representation for complex genomic regions and coding sequences. We assert that the collected updates in GRCh38 make the newer assembly a more robust substrate for comprehensive analyses that will promote our understanding of human biology and advance our efforts to improve health.

Meta-analysis of the difference of medians.

We consider the problem of meta-analyzing two-group studies that report the median of the outcome. Often, these studies are excluded from meta-analysis because there are no well-established statistical methods to pool the difference of medians. To include these studies in meta-analysis, several authors have recently proposed methods to estimate the sample mean and standard deviation from the median, sample size, and several commonly reported measures of spread. Researchers frequently apply these methods to estimate the difference of means and its variance for each primary study and pool the difference of means using inverse variance weighting. In this work, we develop several methods to directly meta-analyze the difference of medians. We conduct a simulation study evaluating the performance of the proposed median-based methods and the competing transformation-based methods. The simulation results show that the median-based methods outperform the transformation-based methods when meta-analyzing studies that report the median of the outcome, especially when the outcome is skewed. Moreover, we illustrate the various methods on a real-life data set.

Streptococcus oralis subsp. dentisani Produces Monolateral Serine-Rich Repeat Protein Fibrils, One of Which Contributes to Saliva Binding via Sialic Acid.

Our studies reveal that the oral colonizer and cause of infective endocarditis Streptococcus oralis subsp. dentisani displays a striking monolateral distribution of surface fibrils. Furthermore, our data suggest that these fibrils impact the structure of adherent bacterial chains. Mutagenesis studies indicate that these fibrils are dependent on three serine-rich repeat proteins (SRRPs), here named fibril-associated protein A (FapA), FapB, and FapC, and that each SRRP forms a different fibril with a distinct distribution. SRRPs are a family of bacterial adhesins that have diverse roles in adhesion and that can bind to different receptors through modular nonrepeat region domains. Amino acid sequence and predicted structural similarity searches using the nonrepeat regions suggested that FapA may contribute to interspecies interactions, that FapA and FapB may contribute to intraspecies interactions, and that FapC may contribute to sialic acid binding. We demonstrate that a fapC mutant was significantly reduced in binding to saliva. We confirmed a role for FapC in sialic acid binding by demonstrating that the parental strain was significantly reduced in adhesion upon addition of a recombinantly expressed, sialic acid-specific, carbohydrate binding module, while the fapC mutant was not reduced. However, mutation of a residue previously shown to be essential for sialic acid binding did not decrease bacterial adhesion, leaving the precise mechanism of FapC-mediated adhesion to sialic acid to be defined. We also demonstrate that the presence of any one of the SRRPs is sufficient for efficient biofilm formation. Similar structures were observed on all infective endocarditis isolates examined, suggesting that this distribution is a conserved feature of this S. oralis subspecies.