VarSifter is a graphical software tool for desktop computers that allows investigators of varying computational skills to easily and quickly sort, filter, and sift through sequence variation data. A ...variety of filters and a custom query framework allow filtering based on any combination of sample and annotation information. By simplifying visualization and analyses of exome-scale sequence variation data, this program will help bring the power and promise of massively-parallel DNA sequencing to a broader group of researchers.
Availability and Implementation: VarSifter is written in Java, and is freely available in source and binary versions, along with a User Guide, at http://research.nhgri.nih.gov/software/VarSifter/.
Contact:
mullikin@mail.nih.gov
Supplementary Information: Additional figures and methods available online at the journal's website.
IMPORTANCE: Identifying infectious causes of subacute or chronic meningitis can be challenging. Enhanced, unbiased diagnostic approaches are needed. OBJECTIVE: To present a case series of patients ...with diagnostically challenging subacute or chronic meningitis using metagenomic next-generation sequencing (mNGS) of cerebrospinal fluid (CSF) supported by a statistical framework generated from mNGS of control samples from the environment and from patients who were noninfectious. DESIGN, SETTING, AND PARTICIPANTS: In this case series, mNGS data obtained from the CSF of 94 patients with noninfectious neuroinflammatory disorders and from 24 water and reagent control samples were used to develop and implement a weighted scoring metric based on z scores at the species and genus levels for both nucleotide and protein alignments to prioritize and rank the mNGS results. Total RNA was extracted for mNGS from the CSF of 7 participants with subacute or chronic meningitis who were recruited between September 2013 and March 2017 as part of a multicenter study of mNGS pathogen discovery among patients with suspected neuroinflammatory conditions. The neurologic infections identified by mNGS in these 7 participants represented a diverse array of pathogens. The patients were referred from the University of California, San Francisco Medical Center (n = 2), Zuckerberg San Francisco General Hospital and Trauma Center (n = 2), Cleveland Clinic (n = 1), University of Washington (n = 1), and Kaiser Permanente (n = 1). A weighted z score was used to filter out environmental contaminants and facilitate efficient data triage and analysis. MAIN OUTCOMES AND MEASURES: Pathogens identified by mNGS and the ability of a statistical model to prioritize, rank, and simplify mNGS results. RESULTS: The 7 participants ranged in age from 10 to 55 years, and 3 (43%) were female. A parasitic worm (Taenia solium, in 2 participants), a virus (HIV-1), and 4 fungi (Cryptococcus neoformans, Aspergillus oryzae, Histoplasma capsulatum, and Candida dubliniensis) were identified among the 7 participants by using mNGS. Evaluating mNGS data with a weighted z score–based scoring algorithm reduced the reported microbial taxa by a mean of 87% (range, 41%-99%) when taxa with a combined score of 0 or less were removed, effectively separating bona fide pathogen sequences from spurious environmental sequences so that, in each case, the causative pathogen was found within the top 2 scoring microbes identified using the algorithm. CONCLUSIONS AND RELEVANCE: Diverse microbial pathogens were identified by mNGS in the CSF of patients with diagnostically challenging subacute or chronic meningitis, including a case of subarachnoid neurocysticercosis that defied diagnosis for 1 year, the first reported case of CNS vasculitis caused by Aspergillus oryzae, and the fourth reported case of C dubliniensis meningitis. Prioritizing metagenomic data with a scoring algorithm greatly clarified data interpretation and highlighted the problem of attributing biological significance to organisms present in control samples used for metagenomic sequencing studies.
The future of DNA sequencing Green, Eric D; Rubin, Edward M; Olson, Maynard V
Nature,
10/2017, Letnik:
550, Številka:
7675
Journal Article
Recenzirano
Odprti dostop
Prognosticators are typically wrong about which technologies - or, more importantly, which applications - will be the most disruptive. We would probably fare no better in predicting the future of DNA ...sequencing. So instead, we offer a framework for thinking about it. Our central message is that trends in DNA sequencing will be driven by killer applications, not by killer technologies.
The UK is set to pilot genetic sequencing in healthy babies. Genomic screening at appropriate ages could help reduce the burden of genetic disorders, say Leslie Biesecker and colleagues, but David ...Curtis argues that newborns cannot consent and that our most personal data might be misused
Charcot-Marie-Tooth disease type 2D (CMT2D) is a dominantly inherited peripheral neuropathy caused by missense mutations in the glycyl-tRNA synthetase gene (GARS). In addition to GARS, mutations in ...three other tRNA synthetase genes cause similar neuropathies, although the underlying mechanisms are not fully understood. To address this, we generated transgenic mice that ubiquitously over-express wild-type GARS and crossed them to two dominant mouse models of CMT2D to distinguish loss-of-function and gain-of-function mechanisms. Over-expression of wild-type GARS does not improve the neuropathy phenotype in heterozygous Gars mutant mice, as determined by histological, functional, and behavioral tests. Transgenic GARS is able to rescue a pathological point mutation as a homozygote or in complementation tests with a Gars null allele, demonstrating the functionality of the transgene and revealing a recessive loss-of-function component of the point mutation. Missense mutations as transgene-rescued homozygotes or compound heterozygotes have a more severe neuropathy than heterozygotes, indicating that increased dosage of the disease-causing alleles results in a more severe neurological phenotype, even in the presence of a wild-type transgene. We conclude that, although missense mutations of Gars may cause some loss of function, the dominant neuropathy phenotype observed in mice is caused by a dose-dependent gain of function that is not mitigated by over-expression of functional wild-type protein.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Cardiovascular disease is a leading cause of morbidity and mortality in individuals with type 2 diabetes mellitus. These high-risk patients benefit from aggressive risk factor management, with blood ...pressure and low-density lipoprotein-cholesterol treatment, glycemic control, kidney protection, and lifestyle intervention. There are several recommendation and guideline documents across cardiology, endocrinology, nephrology, and general medicine professional societies from the United States and Europe with recommendations for cardiovascular risk reduction in patients with type 2 diabetes mellitus. Although there are some noteworthy differences, particularly in risk stratification, low-density lipoprotein-cholesterol and blood pressure treatment targets, and the use of sodium-glucose cotransporter-2 inhibitors and glucagon-like peptide-1 receptor agonists, overall there is considerable alignment across recommendations from different professional societies.
Physical interactions between genetic elements located throughout the genome play important roles in gene regulation and can be identified with the Chromosome Conformation Capture (3C) methodology. ...3C converts physical chromatin interactions into specific ligation products, which are quantified individually by PCR. Here we present a high-throughput 3C approach, 3C-Carbon Copy (5C), that employs microarrays or quantitative DNA sequencing using 454-technology as detection methods. We applied 5C to analyze a 400-kb region containing the human beta-globin locus and a 100-kb conserved gene desert region. We validated 5C by detection of several previously identified looping interactions in the beta-globin locus. We also identified a new looping interaction in K562 cells between the beta-globin Locus Control Region and the gamma-beta-globin intergenic region. Interestingly, this region has been implicated in the control of developmental globin gene switching. 5C should be widely applicable for large-scale mapping of cis- and trans- interaction networks of genomic elements and for the study of higher-order chromosome structure.
BACKGROUND:Financial burden for patients, providers, and payers can reduce access to physical therapy (PT) after total knee arthroplasty (TKA). The purpose of the present study was to examine the ...effect of a virtual PT program on health-care costs and clinical outcomes as compared with traditional care after TKA.
METHODS:At least 10 days before unilateral TKA, patients from 4 clinical sites were enrolled and randomized 1:1 to the virtual PT program (involving an avatar digitally simulated coach, in-home 3-dimensional biometrics, and telerehabilitation with remote clinician oversight by a physical therapist) or to traditional PT care in the home or outpatient clinic. The primary outcome was total health-care costs for the 12-week post-hospital period. Secondary (noninferiority) outcomes included 6 and 12-week Knee injury and Osteoarthritis Outcome Score (KOOS); 6-week knee extension, knee flexion, and gait speed; and 12-week safety measures (patient-reported falls, pain, and hospital readmissions). All outcomes were analyzed on a modified intent-to-treat basis.
RESULTS:Of 306 patients (mean age, 65 years; 62.5% women) who were randomized from November 2016 to November 2017, 290 had TKA and 287 (including 143 in the virtual PT group and 144 in the usual care group) completed the trial. Virtual PT had lower costs at 12 weeks after discharge than usual care (median, $1,050 compared with $2,805; p < 0.001). Mean costs were $2,745 lower for virtual PT patients. Virtual PT patients had fewer rehospitalizations than the usual care group (12 compared with 30; p = 0.007). Virtual PT was noninferior to usual PT in terms of the KOOS at 6 weeks (difference, 0.77; 90% confidence interval CI, −1.68 to 3.23) and 12 weeks (difference, −2.33; 90% CI, −4.98 to 0.31). Virtual PT was also noninferior to usual care at 6 weeks in terms of knee extension, knee flexion, and gait speed and at 12 weeks in terms of pain and hospital readmissions. Falls were reported by 19.4% of virtual PT patients and 14.6% of usual care patients (difference, 4.83%; 90% CI, −2.60 to 12.25).
CONCLUSIONS:Relative to traditional home or clinic PT, virtual PT with telerehabilitation for skilled clinical oversight significantly lowered 3-month health-care costs after TKA while providing similar effectiveness. These findings have important implications for patients, health systems, and payers. Virtual PT with clinical oversight should be considered for patients managed with TKA.
LEVEL OF EVIDENCE:Therapeutic Level I. See Instructions for Authors for a complete description of levels of evidence.
Indolent T-cell lymphoproliferative disorders of the gastrointestinal tract are rare clonal T-cell diseases that more commonly occur in the intestines and have a protracted clinical course. Different ...immunophenotypic subsets have been described, but the molecular pathogenesis and cell of origin of these lymphocytic proliferations is poorly understood. Hence, we performed targeted next-generation sequencing and comprehensive immunophenotypic analysis of ten indolent T-cell lymphoproliferative disorders of the gastrointestinal tract, which comprised CD4
(n=4), CD8
(n=4), CD4
/CD8
(n=1) and CD4
/CD8
(n=1) cases. Genetic alterations, including recurrent mutations and novel rearrangements, were identified in 8/10 (80%) of these lymphoproliferative disorders. The CD4
, CD4
/CD8
, and CD4
/CD8
cases harbored frequent alterations of JAK-STAT pathway genes (5/6, 82%);
mutations (n=3),
deletion (n=1) and
rearrangement (n=1), and 4/6 (67%) had concomitant mutations in epigenetic modifier genes (
,
,
). Conversely, 2/4 (50%) of the CD8
cases exhibited structural alterations involving the 3' untranslated region of the
gene. Longitudinal genetic analysis revealed stable mutational profiles in 4/5 (80%) cases and acquisition of mutations in one case was a harbinger of disease transformation. The CD4
and CD4
/CD8
lymphoproliferative disorders displayed heterogeneous Th1 (T-bet
), Th2 (GATA3
) or hybrid Th1/Th2 (T-bet
/GATA3
) profiles, while the majority of CD8
disorders and the CD4
/CD8
disease showed a type-2 polarized (GATA3
) effector T-cell (Tc2) phenotype. Additionally, CD103 expression was noted in 2/4 CD8
cases. Our findings provide insights into the pathogenetic bases of indolent T-cell lymphoproliferative disorders of the gastrointestinal tract and confirm the heterogeneous nature of these diseases. Detection of shared and distinct genetic alterations of the JAK-STAT pathway in certain immunophenotypic subsets warrants further mechanistic studies to determine whether therapeutic targeting of this signaling cascade is efficacious for a proportion of patients with these recalcitrant diseases.
The completion of a high-quality, comprehensive sequence of the human genome, in this fiftieth anniversary year of the discovery of the double-helical structure of DNA, is a landmark event. The ...genomic era is now a reality.
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