exRNA Atlas Analysis Reveals Distinct Extracellular RNA Cargo Types and Their Carriers Present across Human Biofluids.

To develop a map of cell-cell communication mediated by extracellular RNA (exRNA), the NIH Extracellular RNA Communication Consortium created the exRNA Atlas resource (https://exrna-atlas.org). The Atlas version 4P1 hosts 5,309 exRNA-seq and exRNA qPCR profiles from 19 studies and a suite of analysis and visualization tools. To analyze variation between profiles, we apply computational deconvolution. The analysis leads to a model with six exRNA cargo types (CT1, CT2, CT3A, CT3B, CT3C, CT4), each detectable in multiple biofluids (serum, plasma, CSF, saliva, urine). Five of the cargo types associate with known vesicular and non-vesicular (lipoprotein and ribonucleoprotein) exRNA carriers. To validate utility of this model, we re-analyze an exercise response study by deconvolution to identify physiologically relevant response pathways that were not detected previously. To enable wide application of this model, as part of the exRNA Atlas resource, we provide tools for deconvolution and analysis of user-provided case-control studies.

Surgical treatment of headshaking by removal of a paracondylar process fragment via modified hyovertebrotomy approach: A detailed anatomical and surgical description in an adult horse.

OBJECTIVE: To describe, in detail, the relevant anatomy and surgical approach to access the paracondylar process (PCP) and report its application in a clinical case of headshaking. ANIMAL: A seven-year-old, mixed breed mare. STUDY DESIGN: Experimental study/case report. METHODS: A seven-year-old mixed breed mare was presented for investigation of acute onset progressing violent headshaking, resulting in the horse falling on multiple occasions. The horse was highly reactive to palpation over the right PCP. Standing computed tomographic (CT) investigation and ultrasonographic examination of the head detected a fracture of the right PCP. Five equine heads of mixed breeds and sizes were dissected to demonstrate the relevant anatomy surrounding the PCP with regard to surgical access. A modified hyovertebrotomy approach was used to remove the fracture fragment under general anesthesia. RESULTS: The anatomy surrounding the PCP was described. The fragment was successfully removed resulting in gradual resolution of clinical signs. The horse recovered well postoperatively and was back into light levels of work with no signs of headshaking present two and a half years following surgery. CONCLUSION: The caudal meningeal artery and vein as well as the glossopharyngeal and hypoglossal nerves are adjacent to the PCP and must be avoided during dissections. The modified hyovertebrotomy approach allows safe surgical access to the PCP. Surgical excision of a PCP fragment can result in complete resolution of clinical signs of headshaking. Computed tomography and ultrasonography are valuable diagnostic tools to identify a fracture of the PCP.

Equine odontogenic tumors: Clinical presentation, CT findings, and outcome in 11 horses.

Odontogenic tumors present as locally invasive, slow growing, firm swellings on the face. They are rare in all species and are characterized histologically by the degree of differentiation and dental tissue of origin. Radiographic appearance is not pathognomonic for these lesions. Computed tomographic (CT) examination allows exact determination of tumor extension and aggressiveness. The objectives of this retrospective, case series study were to describe the clinical presentation, CT characteristics, and outcome in horses with histologically confirmed odontogenic tumors, and to identify imaging features suggestive of individual types of tumors. Four ameloblastomas, two ameloblastic carcinomas, three ameloblastic fibromas, and two complex odontomas were included. All but one complex odontoma presented as a single mass. All tumors were associated with maxillary or mandibular bone expansion, alveolar and cortical bone lysis, and cortical bone thinning. The majority also had cortical bone thickening and periosteal proliferation. All tumors contained some degree of mineral attenuation, although only the complex odontomas contained enamel attenuation allowing differentiation from other types of odontogenic tumors in this study. Ameloblastomas were found to have variable CT characteristics likely due to the sub-groups of ameloblastomas. Both ameloblastic carcinomas contained a mixture of mineralized and soft tissue attenuating material whereas ameloblastic fibromas were mainly composed of soft tissue attenuating material. Computed tomographic characteristics of odontogenic tumors generally indicate that they are expansile, aggressive tumors and can occur in a wide range of ages. Further investigation is needed to elucidate differences between each type of equine odontogenic tumor.

ClinGen Allele Registry links information about genetic variants.

Effective exchange of information about genetic variants is currently hampered by the lack of readily available globally unique variant identifiers that would enable aggregation of information from different sources. The ClinGen Allele Registry addresses this problem by providing (1) globally unique "canonical" variant identifiers (CAids) on demand, either individually or in large batches; (2) access to variant-identifying information in a searchable Registry; (3) links to allele-related records in many commonly used databases; and (4) services for adding links to information about registered variants in external sources. A core element of the Registry is a canonicalization service, implemented using in-memory sequence alignment-based index, which groups variant identifiers denoting the same nucleotide variant and assigns unique and dereferenceable CAids. More than 650 million distinct variants are currently registered, including those from gnomAD, ExAC, dbSNP, and ClinVar, including a small number of variants registered by Registry users. The Registry is accessible both via a web interface and programmatically via well-documented Hypertext Transfer Protocol (HTTP) Representational State Transfer Application Programming Interface (REST-APIs). For programmatic interoperability, the Registry content is accessible in the JavaScript Object Notation for Linked Data (JSON-LD) format. We present several use cases and demonstrate how the linked information may provide raw material for reasoning about variant's pathogenicity.

Spark: a navigational paradigm for genomic data exploration.

Biologists possess the detailed knowledge critical for extracting biological insight from genome-wide data resources, and yet they are increasingly faced with nontrivial computational analysis challenges posed by genome-scale methodologies. To lower this computational barrier, particularly in the early data exploration phases, we have developed an interactive pattern discovery and visualization approach, Spark, designed with epigenomic data in mind. Here we demonstrate Spark's ability to reveal both known and novel epigenetic signatures, including a previously unappreciated binding association between the YY1 transcription factor and the corepressor CTBP2 in human embryonic stem cells.

exRNA-eCLIP intersection analysis reveals a map of extracellular RNA binding proteins and associated RNAs across major human biofluids and carriers.

Although the role of RNA binding proteins (RBPs) in extracellular RNA (exRNA) biology is well established, their exRNA cargo and distribution across biofluids are largely unknown. To address this gap, we extend the exRNA Atlas resource by mapping exRNAs carried by extracellular RBPs (exRBPs). This map was developed through an integrative analysis of ENCODE enhanced crosslinking and immunoprecipitation (eCLIP) data (150 RBPs) and human exRNA profiles (6,930 samples). Computational analysis and experimental validation identified exRBPs in plasma, serum, saliva, urine, cerebrospinal fluid, and cell-culture-conditioned medium. exRBPs carry exRNA transcripts from small non-coding RNA biotypes, including microRNA (miRNA), piRNA, tRNA, small nuclear RNA (snRNA), small nucleolar RNA (snoRNA), Y RNA, and lncRNA, as well as protein-coding mRNA fragments. Computational deconvolution of exRBP RNA cargo reveals associations of exRBPs with extracellular vesicles, lipoproteins, and ribonucleoproteins across human biofluids. Overall, we mapped the distribution of exRBPs across human biofluids, presenting a resource for the community.

An integrative variant analysis suite for whole exome next-generation sequencing data.

BACKGROUND: Whole exome capture sequencing allows researchers to cost-effectively sequence the coding regions of the genome. Although the exome capture sequencing methods have become routine and well established, there is currently a lack of tools specialized for variant calling in this type of data. RESULTS: Using statistical models trained on validated whole-exome capture sequencing data, the Atlas2 Suite is an integrative variant analysis pipeline optimized for variant discovery on all three of the widely used next generation sequencing platforms (SOLiD, Illumina, and Roche 454). The suite employs logistic regression models in conjunction with user-adjustable cutoffs to accurately separate true SNPs and INDELs from sequencing and mapping errors with high sensitivity (96.7%). CONCLUSION: We have implemented the Atlas2 Suite and applied it to 92 whole exome samples from the 1000 Genomes Project. The Atlas2 Suite is available for download at http://sourceforge.net/projects/atlas2/. In addition to a command line version, the suite has been integrated into the Genboree Workbench, allowing biomedical scientists with minimal informatics expertise to remotely call, view, and further analyze variants through a simple web interface. The existing genomic databases displayed via the Genboree browser also streamline the process from variant discovery to functional genomics analysis, resulting in an off-the-shelf toolkit for the broader community.

Atlas2 Cloud: a framework for personal genome analysis in the cloud.

BACKGROUND: Until recently, sequencing has primarily been carried out in large genome centers which have invested heavily in developing the computational infrastructure that enables genomic sequence analysis. The recent advancements in next generation sequencing (NGS) have led to a wide dissemination of sequencing technologies and data, to highly diverse research groups. It is expected that clinical sequencing will become part of diagnostic routines shortly. However, limited accessibility to computational infrastructure and high quality bioinformatic tools, and the demand for personnel skilled in data analysis and interpretation remains a serious bottleneck. To this end, the cloud computing and Software-as-a-Service (SaaS) technologies can help address these issues. RESULTS: We successfully enabled the Atlas2 Cloud pipeline for personal genome analysis on two different cloud service platforms: a community cloud via the Genboree Workbench, and a commercial cloud via the Amazon Web Services using Software-as-a-Service model. We report a case study of personal genome analysis using our Atlas2 Genboree pipeline. We also outline a detailed cost structure for running Atlas2 Amazon on whole exome capture data, providing cost projections in terms of storage, compute and I/O when running Atlas2 Amazon on a large data set. CONCLUSIONS: We find that providing a web interface and an optimized pipeline clearly facilitates usage of cloud computing for personal genome analysis, but for it to be routinely used for large scale projects there needs to be a paradigm shift in the way we develop tools, in standard operating procedures, and in funding mechanisms.

A computed tomographic study of endodontic and apical changes in 81 equine cheek teeth with sagittal fractures.

BACKGROUND: Sagittal fractures of equine cheek teeth are commonly observed during oral examination. There are few reports on the apical and endodontic pathology associated with such fractures seen during computed tomographic (CT) examination. OBJECTIVES: This study aimed to document the prevalence of CT changes indicative of apical disease in equine cheek teeth, which have suffered a sagittal fracture involving the clinical ± reserve crown. STUDY DESIGN: This study is a retrospective case series. METHODS: CT examinations of equine heads with sagittal fractures of cheek teeth present were reviewed: 81 teeth from 49 horses were identified to have a sagittal cheek tooth fracture. The images were evaluated for apical pathology including gas (in the endodontic system and periapically), widened periodontal space, periapical sclerosis, apical clubbing, cementoma/hypercementosis, lamina dura loss, associated sinusitis and sinus mucosal swelling. An apical infection grading system was created to give each tooth a score. Hounsfield units were used to measure the density of the endodontic, apical and periapical regions. The fracture length ratio was recorded. Statistical analysis was performed using a generalised estimating equation to evaluate predictors of apical infection and associations between clinical signs and CT abnormalities. RESULTS: Eighty-seven sagittal fractures (56 buccal, 17 palatal/lingual and 14 midline) from 81 teeth were recorded (74 maxillary and 7 mandibular). Apical infection was diagnosed in 73% (37/51, P = .05) of buccal, 55% (6/11, P = .07) of palatal/lingual, 100% (13/13) of midline, 100% (6/6) of multiple fractures and 96% (23/24, P = .008) of fractures involving infundibula. There was no significant relationship between apical infection and the presence of clinical signs associated with dental pathology (P = .4). There was no significant association between fracture length ratio and apical infection (P = 1.0). Midline sagittal fractures were significantly associated with sinusitis when compared with all other maxillary fractures (odds ratio [OR] 5.92, 95% confidence interval [CI] 1.67-20.83, P = .006). Loss of the lamina dura was not significantly associated with apical infection (P = .5). MAIN LIMITATIONS: There is a maxillary cheek tooth bias in the data set and the subjective grading system. CONCLUSIONS: A large proportion of fractured cheek teeth have evidence of apical infection on CT examination and therefore warrant treatment.

Diagnosis, management, and outcome in 19 horses with deltoid tuberosity fractures.

OBJECTIVE: To describe the diagnosis and treatment of fractures of the deltoid tuberosity. STUDY DESIGN: Case series. METHODS: Medical records (1992-2009) of 19 horses with radiographic confirmation of deltoid tuberosity fractures were reviewed. Data retrieved included signalment, clinical and diagnostic imaging findings, and treatment. Outcome was determined by telephone questionnaire of owners and referring veterinarians. RESULTS: Most horses were markedly lame on admission and 53% had reduced protraction of the affected limb. All fractures were identified on a cranio45° medial-caudolateral oblique projection; however, only 32% (6 horses) were detected on a mediolateral projection whereas 86% were evident ultrasonographically. Treatment by local wound care and stall rest resulted in return to athletic function without lameness for 13 of 14 horses that had follow-up. CONCLUSIONS: A cranio45° medial-caudolateral oblique radiographic view was better than a mediolateral projection for identification of deltoid tuberosity fractures. Ultrasonographic detection of fractures was similar except when gas accumulation obscured the fracture site. Deltoid tuberosity fractures can cause severe lameness but can be treated successfully with conservative management.

The effect of tape type, taping method and tape storage temperature on the retrieval rate of fibres from various surfaces: An example of data generation and analysis to facilitate trace evidence recovery validation and optimisation.

This paper aspires to assist those tasked with data generation and analysis for the purpose of the validation and/or optimisation of trace evidence recovery. It does so via a detailed report of the authors' approach to this problem in the context of target fibre retrieval using self-adhesive tapes. Textile fibres can provide valuable evidence at both source and activity levels. This ability stems from their near ubiquity in the man-made environment, their potential for high levels of discrimination (especially when found in combination) and their reproducible transfer and persistence behaviours. To realise this value for the criminal justice system, it is vital that police forces and forensic providers are collectively able to search for, recover and analyse fibres found at crime scenes and correctly evaluate their evidential value. ISO accreditation provides quality assurance for such activities. The work reported in this paper was part of a study to validate crime scene fibre retrieval processes for the purposes of ISO17020 accreditation. However, it is hoped that it will be of assistance to those wishing to validate and/or optimise forensic fibre recovery whether at the crime scene or in the laboratory. Further, the methods described may be of value to those who need to validate and/or optimise the recovery of other types of trace evidence. This paper outlines a series of experiments that investigated the effect of four factors on the rate at which target fibres could be recovered from surfaces by tape lifting. The factors were tape type (with two levels, namely: J-LAR and Crystal Tabs), tape storage temperature (three levels: -5 °C, room temperature [19 ±â€¯1 °C] and 35 °C), taping method (two levels: zonal and one-to-one) and surface (12 levels: each being a surface type commonly encountered at crime scenes). This resulted in 144 unique experimental conditions. For each of these, five repeat fibre recovery rate determinations were carried out, generating 720 data points. All surfaces were clean and dry prior to target fibres being transferred and recovered. In all cases, the tapes were applied to the surfaces at 19 ±â€¯1 °C. These experiments showed that the surfaces can be divided into three stable clusters based on the median and interquartile range of the fibre retrieval rate achieved from each of them. Also, they showed that, in terms of the proportion of the target fibres retrieved, typically and setting aside interaction effects: However, notably, a good degree of between-condition overlap was also apparent in the data. To understand this, a four-way factorial ANOVA model was built which revealed significant and substantive effects for all four main effects and for 10 of the 11 interactions. Importantly, the four-way interaction term was amongst those found to be significant. The interplay between the effects of the four factors was analysed by means of simple effects tests and pairwise contrasts. Tables and interactive parallel coordinate plots have been created. Using these it can easily be seen which of any given pair of levels of each of the four factors resulted in the higher fibre retrieval rate under any one of the unique conditions of the study, and the effect size and statistical significance of this observation. Qualitative evaluations of the effect of tape storage temperatures on tape pliability and its propensity to tear in use were also made.

ClinGen Pathogenicity Calculator: a configurable system for assessing pathogenicity of genetic variants.

BACKGROUND: The success of the clinical use of sequencing based tests (from single gene to genomes) depends on the accuracy and consistency of variant interpretation. Aiming to improve the interpretation process through practice guidelines, the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP) have published standards and guidelines for the interpretation of sequence variants. However, manual application of the guidelines is tedious and prone to human error. Web-based tools and software systems may not only address this problem but also document reasoning and supporting evidence, thus enabling transparency of evidence-based reasoning and resolution of discordant interpretations. RESULTS: In this report, we describe the design, implementation, and initial testing of the Clinical Genome Resource (ClinGen) Pathogenicity Calculator, a configurable system and web service for the assessment of pathogenicity of Mendelian germline sequence variants. The system allows users to enter the applicable ACMG/AMP-style evidence tags for a specific allele with links to supporting data for each tag and generate guideline-based pathogenicity assessment for the allele. Through automation and comprehensive documentation of evidence codes, the system facilitates more accurate application of the ACMG/AMP guidelines, improves standardization in variant classification, and facilitates collaborative resolution of discordances. The rules of reasoning are configurable with gene-specific or disease-specific guideline variations (e.g. cardiomyopathy-specific frequency thresholds and functional assays). The software is modular, equipped with robust application program interfaces (APIs), and available under a free open source license and as a cloud-hosted web service, thus facilitating both stand-alone use and integration with existing variant curation and interpretation systems. The Pathogenicity Calculator is accessible at http://calculator.clinicalgenome.org . CONCLUSIONS: By enabling evidence-based reasoning about the pathogenicity of genetic variants and by documenting supporting evidence, the Calculator contributes toward the creation of a knowledge commons and more accurate interpretation of sequence variants in research and clinical care.

Epigenomic footprints across 111 reference epigenomes reveal tissue-specific epigenetic regulation of lincRNAs.

Tissue-specific expression of lincRNAs suggests developmental and cell-type-specific functions, yet tissue specificity was established for only a small fraction of lincRNAs. Here, by analysing 111 reference epigenomes from the NIH Roadmap Epigenomics project, we determine tissue-specific epigenetic regulation for 3,753 (69% examined) lincRNAs, with 54% active in one of the 14 cell/tissue clusters and an additional 15% in two or three clusters. A larger fraction of lincRNA TSSs is marked in a tissue-specific manner by H3K4me1 than by H3K4me3. The tissue-specific lincRNAs are strongly linked to tissue-specific pathways and undergo distinct chromatin state transitions during cellular differentiation. Polycomb-regulated lincRNAs reside in the bivalent state in embryonic stem cells and many of them undergo H3K27me3-mediated silencing at early stages of differentiation. The exquisitely tissue-specific epigenetic regulation of lincRNAs and the assignment of a majority of them to specific tissue types will inform future studies of this newly discovered class of genes.

Distribution of injected technetium(99m)-labeled mesenchymal stem cells in horses with naturally occurring tendinopathy.

This study aimed to investigate immediate cell survival and distribution following different administration routes of mesenchymal stem cells (MSCs) into naturally occurring tendon injuries. Ten million MSCs, labeled with technetium-99m hexamethylpropyleneamine oxime, were implanted into 13 horses with naturally occurring tendon or ligament injuries intra-lesionally, intravenously and by regional perfusion, and traced for up to 48 h using planar gamma scintigraphy. Labeling efficiencies varied between 1.8% and 18.5% (mean 9.3%). Cells were retained in the damaged area after intra-lesional administration but only 24% of cells were still present within the tendon after 24 h. After intravenous injection, cells largely distributed to the lung fields, with no detectable cells in the tendon lesions. Significant labeling of the tendon lesions was observed in 11/12 horses following regional perfusion but at a lower level to intra-lesional injection. The highest cell numbers were retained after intra-lesional injection, although with considerable cell loss, while regional perfusion may be a viable alternative for MSC delivery. Cells did not "home" to damaged tendon in large numbers after intravenous administration. Cells were detected in the lungs most frequently after intravascular administration, although with no adverse effects. Low cell retention has important implications for designing effective clinical therapies for human clinical use.

Pooled genomic indexing of rhesus macaque.

Pooled genomic indexing (PGI) is a method for mapping collections of bacterial artificial chromosome (BAC) clones between species by using a combination of clone pooling and DNA sequencing. PGI has been used to map a total of 3858 BAC clones covering approximately 24% of the rhesus macaque (Macaca mulatta) genome onto 4178 homologous loci in the human genome. A number of intrachromosomal rearrangements were detected by mapping multiple segments within the individual rhesus BACs onto multiple disjoined loci in the human genome. Transversal pooling designs involving shuffled BAC arrays were employed for robust mapping even with modest DNA sequence read coverage. A further innovation, short-tag pooled genomic indexing (ST-PGI), was also introduced to further improve the economy of mapping by sequencing multiple, short, mapable tags within a single sequencing reaction.

Pash: efficient genome-scale sequence anchoring by Positional Hashing.

Pash is a computer program for efficient, parallel, all-against-all comparison of very long DNA sequences. Pash implements Positional Hashing, a novel parallelizable method for sequence comparison based on k-mer representation of sequences. The Positional Hashing method breaks the comparison problem in a unique way that avoids the quadratic penalty encountered with other sensitive methods and confers inherent low-level parallelism. Furthermore, Positional Hashing allows one to readily and predictably trade between sensitivity and speed. In a simulated comparison task, anchoring computationally mutated reads onto a genome, the sensitivity of Pash was equal to or greater than that of BLAST and BLAT, with Pash outperforming these programs as the reads became shorter and less similar to the genome. Using modest computing resources, we employed Pash for two large-scale sequence comparison tasks: comparison of three mammalian genomes, and anchoring millions of chimpanzee whole-genome shotgun sequencing reads onto the human genome. The results of these comparisons by Pash agree with those computed by other methods that use more than an order of magnitude more computing resources. These results confirm the sensitivity of Positional Hashing.