TreeToReads - a pipeline for simulating raw reads from phylogenies.

BackgroundUsing phylogenomic analysis tools for tracking pathogens has become standard practice in academia, public health agencies, and large industries. Using the same raw read genomic data as input, there are several different approaches being used to infer phylogenetic tree. These include many different SNP pipelines, wgMLST approaches, k-mer algorithms, whole genome alignment and others; each of these has advantages and disadvantages, some have been extensively validated, some are faster, some have higher resolution. A few of these analysis approaches are well-integrated into the regulatory process of US Federal agencies (e.g. the FDA's SNP pipeline for tracking foodborne pathogens). However, despite extensive validation on benchmark datasets and comparison with other pipelines, we lack methods for fully exploring the effects of multiple parameter values in each pipeline that can potentially have an effect on whether the correct phylogenetic tree is recovered.ResultsTo resolve this problem, we offer a program, TreeToReads, which can generate raw read data from mutated genomes simulated under a known phylogeny. This simulation pipeline allows direct comparisons of simulated and observed data in a controlled environment. At each step of these simulations, researchers can vary parameters of interest (e.g., input tree topology, amount of sequence divergence, rate of indels, read coverage, distance of reference genome, etc) to assess the effects of various parameter values on correctly calling SNPs and reconstructing an accurate tree.ConclusionsSuch critical assessments of the accuracy and robustness of analytical pipelines are essential to progress in both research and applied settings

Duplex Ultrasonography to Predict Internal Carotid Artery Stenoses Exceeding 50% and 70% as Defined by NASCET: The Need for Multiple Criteria

Carotid duplex scanning is being used more frequently as the sole preoperative diagnostic imaging modality for patients considered candidates for carotid endarterectomy. The North American Symptomatic Carotid Endarterectomy Trial (NASCET) has demonstrated the benefit of surgical treatment in patients with carotid stenoses exceeding 70%. The purpose of this study was to determine duplex criteria that accurately predict carotid stenoses exceeding 50% and 70% as defined by NASCET arteriographic criteria. One hundred forty-one patients (264 carotid arteries) considered surgical candidates were prospectively studied over a 2-year period by use of both duplex scanning and digital subtraction cerebral arteriography. Carotid artery stenosis was determined by a single radiologist using NASCET arteriographic criteria. Peak systolic velocity (PSV) and enddiastolic velocity (EDV) were measured in the internal carotid (ICA) and common carotid (CCA) arteries by use of duplex scanning. ICA/CCA velocity ratios were calculated for PSV and EDV. Sensitivity, specificity, positive and negative predictive values, and accuracy were calculated. PSVICA/CCA provided the highest sensitivity, and EDVICA the highest specificity in this study. Arteriographic stenoses exceeding 50% and 70% were reliably predicted with use of these duplex criteria. It is concluded that duplex criteria can predict carotid stenoses exceeding 50% and 70% as defined by NASCET arteriographic criteria. These criteria should be independently validated by other vascular laboratories.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/68440/2/10.1177_153857449903300508.pd

Proficiency testing for bacterial whole genome sequencing: an end-user survey of current capabilities, requirements and priorities

The advent of next-generation sequencing (NGS) has revolutionised public health microbiology. Given the potential impact of NGS, it is paramount to ensure standardisation of ‘wet’ laboratory and bioinformatic protocols and promote comparability of methods employed by different laboratories and their outputs. Therefore, one of the ambitious goals of the Global Microbial Identifier (GMI) initiative (http://www.globalmicrobialidentifier.org/) has been to establish a mechanism for inter-laboratory NGS proficiency testing (PT). This report presents findings from the survey recently conducted by Working Group 4 among GMI members in order to ascertain NGS end-use requirements and attitudes towards NGS PT. The survey identified the high professional diversity of laboratories engaged in NGS-based public health projects and the wide range of capabilities within institutions, at a notable range of costs. The priority pathogens reported by respondents reflected the key drivers for NGS use (high burden disease and ‘high profile’ pathogens). The performance of and participation in PT was perceived as important by most respondents. The wide range of sequencing and bioinformatics practices reported by end-users highlights the importance of standardisation and harmonisation of NGS in public health and underpins the use of PT as a means to assuring quality. The findings of this survey will guide the design of the GMI PT program in relation to the spectrum of pathogens included, testing frequency and volume as well as technical requirements. The PT program for external quality assurance will evolve and inform the introduction of NGS into clinical and public health microbiology practice in the post-genomic era. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12879-015-0902-3) contains supplementary material, which is available to authorized users

Large-scale functional inference for skin-expressing lncRNAs using expression and sequence information

Long noncoding RNAs (lncRNAs) regulate the expression of protein-coding genes and have been shown to play important roles in inflammatory skin diseases. However, we still have limited understanding of the functional impact of lncRNAs in skin, partly due to their tissue specificity and lower expression levels compared with protein-coding genes. We compiled a comprehensive list of 18,517 lncRNAs from different sources and studied their expression profiles in 834 RNA-Seq samples from multiple inflammatory skin conditions and cytokine-stimulated keratinocytes. Applying a balanced random forest to predict involvement in biological functions, we achieved a median AUROC of 0.79 in 10-fold cross-validation, identifying significant DNA binding domains (DBDs) for 39 lncRNAs. G18244, a skin-expressing lncRNA predicted for IL-4/IL-13 signaling in keratinocytes, was highly correlated in expression with F13A1, a protein-coding gene involved in macrophage regulation, and we further identified a significant DBD in F13A1 for G18244. Reflecting clinical implications, AC090198.1 (predicted for IL-17 pathway) and AC005332.6 (predicted for IFN-γ pathway) had significant negative correlation with the SCORAD metric for atopic dermatitis. We also utilized single-cell RNA and spatial sequencing data to validate cell type specificity. Our research demonstrates lncRNAs have important immunological roles and can help prioritize their impact on inflammatory skin diseases.</p

Transferability and Fine Mapping of Type 2 Diabetes Loci in African Americans: The Candidate Gene Association Resource Plus Study

Type 2 diabetes (T2D) disproportionally affects African Americans (AfA) but, to date, genetic variants identified from genome-wide association studies (GWAS) are primarily from European and Asian populations. We examined the single nucleotide polymorphism (SNP) and locus transferability of 40 reported T2D loci in six AfA GWAS consisting of 2,806 T2D case subjects with or without end-stage renal disease and 4,265 control subjects from the Candidate Gene Association Resource Plus Study. Our results revealed that seven index SNPs at the TCF7L2, KLF14, KCNQ1, ADCY5, CDKAL1, JAZF1, and GCKR loci were significantly associated with T2D (P < 0.05). The strongest association was observed at TCF7L2 rs7903146 (odds ratio [OR] 1.30; P = 6.86 × 10−8). Locus-wide analysis demonstrated significant associations (Pemp < 0.05) at regional best SNPs in the TCF7L2, KLF14, and HMGA2 loci as well as suggestive signals in KCNQ1 after correction for the effective number of SNPs at each locus. Of these loci, the regional best SNPs were in differential linkage disequilibrium (LD) with the index and adjacent SNPs. Our findings suggest that some loci discovered in prior reports affect T2D susceptibility in AfA with similar effect sizes. The reduced and differential LD pattern in AfA compared with European and Asian populations may facilitate fine mapping of causal variants at loci shared across populations

Thrombosis of the vasa vasorum of the large and medium size pulmonary artery and vein leads to pulmonary thromboembolism in COVID-19

The vasa vasorum of the large pulmonary vessels is involved in the pathology of COVID-19. This specialized microvasculature plays a major role in the biology and pathology of the pulmonary vessel walls. We have evidence that thrombosis of the vasa vasorum of the large and medium-sized pulmonary vessels during severe COVID-19 causes ischemia and subsequent death of the pulmonary vasculature endothelium. Subsequent release of thrombi from the vasa interna into the pulmonary circulation and pulmonary embolism generated at the ischemic pulmonary vascular endothelium site, are the central pathophysiological mechanisms in COVID-19 responsible for pulmonary thromboembolism. The thrombosis of the vasa vasorum of the large and medium-sized pulmonary vessels is an internal event leading to pulmonary thromboembolism in COVID-19

Opportunities to integrate new approaches in genetic toxicology: An ILSI-HESI workshop report

Genetic toxicity tests currently used to identify and characterize potential human mutagens and carcinogens rely on measurements of primary DNA damage, gene mutation, and chromosome damage in vitro and in rodents. The International Life Sciences Institute Health and Environmental Sciences Institute (ILSI-HESI) Committee on the Relevance and Follow-up of Positive Results in In Vitro Genetic Toxicity Testing held an April 2012 Workshop in Washington, DC, to consider the impact of new understanding of biology and new technologies on the identification and characterization of genotoxic substances, and to identify new approaches to inform more accurate human risk assessment for genetic and carcinogenic effects. Workshop organizers and speakers were from industry, academe, and government. The Workshop focused on biological effects and technologies that would potentially yield the most useful information for evaluating human risk of genetic damage. Also addressed was the impact that improved understanding of biology and availability of new techniques might have on genetic toxicology practices. Workshop topics included (1) alternative experimental models to improve genetic toxicity testing, (2) Biomarkers of epigenetic changes and their applicability to genetic toxicology, and (3) new technologies and approaches. The ability of these new tests and technologies to be developed into tests to identify and characterize genotoxic agents; to serve as a bridge between in vitro and in vivo rodent, or preferably human, data; or to be used to provide dose response information for quantitative risk assessment was also addressed. A summary of the workshop and links to the scientific presentations are provided.International Life Sciences Institute/Health and Environmental Sciences Institute Committe