Whole Genome Sequencing in the Clinical Laboratory

2012 ◽  
pp. 67-78
Author(s):  
Tina Hambuch ◽  
Brad Sickler ◽  
Arnold Liao ◽  
Suneer Jain ◽  
Philip D. Cotter
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Clinical Laboratory ◽  
Whole Genome

scholarly journals Precision medicine integrating whole-genome sequencing, comprehensive metabolomics, and advanced imaging

2020 ◽  
Vol 117 (6) ◽  
pp. 3053-3062 ◽  
Author(s):  
Ying-Chen Claire Hou ◽  
Hung-Chun Yu ◽  
Rick Martin ◽  
Elizabeth T. Cirulli ◽  
Natalie M. Schenker-Ahmed ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Precision Medicine ◽  
Genome Sequencing ◽  
Clinical Laboratory ◽  
Descriptive Analysis ◽  
Disease Diagnosis ◽  
Whole Genome ◽  
Advanced Imaging ◽  
Family Medical History ◽  
Deep Phenotyping

Genome sequencing has established clinical utility for rare disease diagnosis. While increasing numbers of individuals have undergone elective genome sequencing, a comprehensive study surveying genome-wide disease-associated genes in adults with deep phenotyping has not been reported. Here we report the results of a 3-y precision medicine study with a goal to integrate whole-genome sequencing with deep phenotyping. A cohort of 1,190 adult participants (402 female [33.8%]; mean age, 54 y [range 20 to 89+]; 70.6% European) had whole-genome sequencing, and were deeply phenotyped using metabolomics, advanced imaging, and clinical laboratory tests in addition to family/medical history. Of 1,190 adults, 206 (17.3%) had at least 1 genetic variant with pathogenic (P) or likely pathogenic (LP) assessment that suggests a predisposition of genetic risk. A multidisciplinary clinical team reviewed all reportable findings for the assessment of genotype and phenotype associations, and 137 (11.5%) had genotype and phenotype associations. A high percentage of genotype and phenotype associations (>75%) was observed for dyslipidemia (n = 24), cardiomyopathy, arrhythmia, and other cardiac diseases (n = 42), and diabetes and endocrine diseases (n = 17). A lack of genotype and phenotype associations, a potential burden for patient care, was observed in 69 (5.8%) individuals with P/LP variants. Genomics and metabolomics associations identified 61 (5.1%) heterozygotes with phenotype manifestations affecting serum metabolite levels in amino acid, lipid and cofactor, and vitamin pathways. Our descriptive analysis provides results on the integration of whole-genome sequencing and deep phenotyping for clinical assessments in adults.

scholarly journals The Brazilian Rare Genomes Project: validation of whole genome sequencing for rare diseases diagnosis

2021 ◽  
Author(s):  
Antonio Victor Campos Coelho ◽  
Bruna Mascaro Cordeiro de Azevedo ◽  
Danielle Ribeiro Lucon ◽  
Maria Soares Nobrega ◽  
Rodrigo de Souza Reis ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Rare Diseases ◽  
Clinical Laboratory ◽  
Genomic Medicine ◽  
Uniparental Disomy ◽  
Public Healthcare ◽  
Whole Genome ◽  
Single Nucleotide Variants ◽  
F Measure

Rare diseases affect 3.2 to 13.2 million individuals in Brazil. The Brazilian Rare Genomes Project is envisioned to further the implementation of genomic medicine into the Brazilian public healthcare system. Here we report the results of the validation of a whole genome sequencing (WGS) procedure for implementation in a clinical laboratory. In addition, we report data quality for the first 1,200 real world patients sequenced. For the validation, we sequenced a well characterized group of 76 samples, including seven gold standard genomes, using a PCR-free WGS protocol on Illumina Novaseq 6000 equipment. We compared the observed variant calls with their expected calls, observing good concordance for single nucleotide variants (SNVs; mean F-measure = 99.82%) and indels (mean F-measure = 99.57%). Copy number variants and structural variants events detection performances were as expected (F-measures 96.6% and 90.3%, respectively). Our protocol presented excellent intra- and inter-assay reproducibility, with coefficients of variation ranging between 0.03% and 0.20% and 0.02% and 0.09%, respectively. Limitations of the procedure include the inability to confidently detect variants such as uniparental disomy, balanced translocations, repeat expansion variants and low-level mosaicism. In summary, the observed performance of the test was in accordance with that seen in the best centers worldwide. The Rare Genomes Project is an important initiative to improve Brazil's general population access to the innovative WGS technology which has the potential to reduce the time until diagnosis of rare diseases, bringing pivotal improvements for the quality of life of the affected individuals.

scholarly journals Validation and Implementation of Clinical Laboratory Improvements Act-Compliant Whole-Genome Sequencing in the Public Health Microbiology Laboratory

2017 ◽  
Vol 55 (8) ◽  
pp. 2502-2520 ◽  
Author(s):  
Varvara K. Kozyreva ◽  
Chau-Linda Truong ◽  
Alexander L. Greninger ◽  
John Crandall ◽  
Rituparna Mukhopadhyay ◽  
...  
Keyword(s):  
Public Health ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Clinical Laboratory ◽  
Limit Of Detection ◽  
Control Measures ◽  
Whole Genome ◽  
Content Type ◽  
Specificity And Sensitivity ◽  
Performance Specifications

ABSTRACT Public health microbiology laboratories (PHLs) are on the cusp of unprecedented improvements in pathogen identification, antibiotic resistance detection, and outbreak investigation by using whole-genome sequencing (WGS). However, considerable challenges remain due to the lack of common standards. Here, we describe the validation of WGS on the Illumina platform for routine use in PHLs according to Clinical Laboratory Improvements Act (CLIA) guidelines for laboratory-developed tests (LDTs). We developed a validation panel comprising 10 Enterobacteriaceae isolates, 5 Gram-positive cocci, 5 Gram-negative nonfermenting species, 9 Mycobacterium tuberculosis isolates, and 5 miscellaneous bacteria. The genome coverage range was 15.71× to 216.4× (average, 79.72×; median, 71.55×); the limit of detection (LOD) for single nucleotide polymorphisms (SNPs) was 60×. The accuracy, reproducibility, and repeatability of base calling were >99.9%. The accuracy of phylogenetic analysis was 100%. The specificity and sensitivity inferred from multilocus sequence typing (MLST) and genome-wide SNP-based phylogenetic assays were 100%. The following objectives were accomplished: (i) the establishment of the performance specifications for WGS applications in PHLs according to CLIA guidelines, (ii) the development of quality assurance and quality control measures, (iii) the development of a reporting format for end users with or without WGS expertise, (iv) the availability of a validation set of microorganisms, and (v) the creation of a modular template for the validation of WGS processes in PHLs. The validation panel, sequencing analytics, and raw sequences could facilitate multilaboratory comparisons of WGS data. Additionally, the WGS performance specifications and modular template are adaptable for the validation of other platforms and reagent kits.

scholarly journals Lipid A-Ara4N as an alternate pathway for (colistin) resistance in Klebsiella pneumonia isolates in Pakistan

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Kiran Iqbal Masood ◽  
Seema Umar ◽  
Zahra Hasan ◽  
Joveria Farooqi ◽  
Safina Abdul Razzak ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Lipid A ◽  
Clinical Laboratory ◽  
Laboratory Strain ◽  
Klebsiella Pneumonia ◽  
Whole Genome ◽  
Rt Pcr ◽  
Colistin Resistance ◽  
Alternate Pathway

Abstract Objectives This study aimed to explore mechanism of colistin resistance amongst Klebsiella pneumoniae isolates through plasmid mediated mcr-1 gene in Pakistan. Carbapenem and Colistin resistant K. pneumoniae isolates (n  = 34) stored at − 80 °C as part of the Aga Khan University Clinical Laboratory strain bank were randomly selected and subjected to mcr-1 gene PCR. To investigate mechanisms of resistance, other than plasmid mediated mcr-1 gene, whole genome sequencing was performed on 8 clinical isolates, including 6 with colistin resistance (MIC  >  4 μg/ml) and 2 with intermediate resistance to colistin (MIC  >  2 μg/ml). Results RT-PCR conducted revealed absence of mcr-1 gene in all isolates tested. Whole genome sequencing results revealed modifications in Lipid A-Ara4N pathway. Modifications in Lipid A-Ara4N pathway were detected in ArnA_ DH/FT, UgdH, ArnC and ArnT genes. Mutation in ArnA_ DH/FT gene were detected in S3, S5, S6 and S7 isolates. UgdH gene modifications were found in all isolates except S3, mutations in ArnC were present in all except S1, S2 and S8 and ArnT were detected in all except S4 and S7. In the absence of known mutations linked with colistin resistance, lipid pathway modifications may possibly explain the phenotype resistance to colistin, but this needs further exploration.

scholarly journals Integrating Whole-Genome Sequencing in Clinical Genetics: A Novel Disruptive Structural Rearrangement Identified in the Dystrophin Gene (DMD)

2021 ◽  
Vol 23 (1) ◽  
pp. 59
Author(s):  
Ana Gonçalves ◽  
Ana Fortuna ◽  
Yavuz Ariyurek ◽  
Márcia E. Oliveira ◽  
Goreti Nadais ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Clinical Laboratory ◽  
Dystrophin Gene ◽  
Genetic Alterations ◽  
Molecular Techniques ◽  
Clinical Genetics ◽  
Whole Genome ◽  
Transcript Analysis ◽  
Mild Intellectual Disability

While in most patients the identification of genetic alterations causing dystrophinopathies is a relatively straightforward task, a significant number require genomic and transcriptomic approaches that go beyond a routine diagnostic set-up. In this work, we present a Becker Muscular Dystrophy patient with elevated creatinine kinase levels, progressive muscle weakness, mild intellectual disability and a muscle biopsy showing dystrophic features and irregular dystrophin labelling. Routine molecular techniques (Southern-blot analysis, multiplex PCR, MLPA and genomic DNA sequencing) failed to detect a defect in the DMD gene. Muscle DMD transcript analysis (RT-PCR and cDNA-MLPA) showed the absence of exons 75 to 79, seen to be present at the genomic level. These results prompted the application of low-coverage linked-read whole-genome sequencing (WGS), revealing a possible rearrangement involving DMD intron 74 and a region located upstream of the PRDX4 gene. Breakpoint PCR and Sanger sequencing confirmed the presence of a ~8 Mb genomic inversion. Aberrant DMD transcripts were subsequently identified, some of which contained segments from the region upstream of PRDX4. Besides expanding the mutational spectrum of the disorder, this study reinforces the importance of transcript analysis in the diagnosis of dystrophinopathies and shows how WGS has a legitimate role in clinical laboratory genetics.

scholarly journals Isoniazid resistance in Mycobacterium tuberculosis is a heterogeneous phenotype comprised of overlapping MIC distributions with different underlying resistance mechanisms

2019 ◽  
Author(s):  
Arash Ghodousi ◽  
Elisa Tagliani ◽  
Eranga Karunaratne ◽  
Stefan Niemann ◽  
Jennifer Perera ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Clinical Laboratory ◽  
Resistance Mechanisms ◽  
Whole Genome ◽  
Isoniazid Resistance ◽  
Low Level ◽  
Promoter Mutations ◽  
Level Resistance

AbstractMIC testing using the BACTEC 960 MGIT system of 70 phylogenetically diverse, isoniazid-resistant clinical strains of Mycobacterium tuberculosis revealed a complex pattern of overlapping MIC distributions. Whole-genome sequencing could explain most of the level of resistance observed. The MIC distribution of strains with only inhA promoter mutations was split by the current concentration that is endorsed by the Clinical Laboratory Standards Institute to detect low-level resistance to isoniazid and is therefore likely not optimally set.

scholarly journals Whole genome sequencing in the Middle Eastern Qatari population identifies genetic associations with 45 clinically relevant traits

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Gaurav Thareja ◽  
◽  
Yasser Al-Sarraj ◽  
Aziz Belkadi ◽  
Maryam Almotawa ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Genome Wide Association Study ◽  
Clinical Laboratory ◽  
Medical Decision ◽  
Future Application ◽  
Whole Genome ◽  
Genome Wide ◽  
A Genome ◽  
Polygenic Scores

AbstractClinical laboratory tests play a pivotal role in medical decision making, but little is known about their genetic variability between populations. We report a genome-wide association study with 45 clinically relevant traits from the population of Qatar using a whole genome sequencing approach in a discovery set of 6218 individuals and replication in 7768 subjects. Trait heritability is more similar between Qatari and European populations (r = 0.81) than with Africans (r = 0.44). We identify 281 distinct variant-trait-associations at genome wide significance that replicate known associations. Allele frequencies for replicated loci show higher correlations with European (r = 0.94) than with African (r = 0.85) or Japanese (r = 0.80) populations. We find differences in linkage disequilibrium patterns and in effect sizes of the replicated loci compared to previous reports. We also report 17 novel and Qatari-predominate signals providing insights into the biological pathways regulating these traits. We observe that European-derived polygenic scores (PGS) have reduced predictive performance in the Qatari population which could have implications for the translation of PGS between populations and their future application in precision medicine.

Utility of Whole Genome Sequencing in diagnosing complex disorders: lesson from renal tubular disorders

2018 ◽  
Author(s):  
Mark Stevenson ◽  
Alistair T Pagnamenta ◽  
Heather G Mack ◽  
Judith A Savige ◽  
Kate E Lines ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Whole Genome ◽  
Complex Disorders ◽  
Tubular Disorders ◽  
Renal Tubular Disorders ◽  
Renal Tubular

0306 Exploring the feasibility of using copy number variants as genetic markers through large-scale whole genome sequencing experiments

2016 ◽  
Vol 94 (suppl_5) ◽  
pp. 146-146
Author(s):  
D. M. Bickhart ◽  
L. Xu ◽  
J. L. Hutchison ◽  
J. B. Cole ◽  
D. J. Null ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genetic Markers ◽  
Genome Sequencing ◽  
Copy Number ◽  
Large Scale ◽  
Copy Number Variants ◽  
Whole Genome
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