scholarly journals Complete assembly of novel environmental bacterial genomes by MinIONTM sequencing

2015 ◽  
Author(s):  
Daniel J Turner ◽  
Xiaoguang Dai ◽  
Simon Mayes ◽  
Sissel Juul
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Genomic Analysis ◽  
Whole Genome ◽  
Bacterial Isolates ◽  
Bacterial Genomes ◽  
Prokaryotic Genomes ◽  
Sequencing Process

In this study, we adapt a protocol for the growth of previously uncultured environmental bacterial isolates, to make it compatible with whole genome sequencing. We demonstrate that in combination with the MinION sequencing device, complete assemblies can be derived, allowing genomic comparisons to be made. This approach allows rapid, inexpensive and straightforward discovery, and genomic analysis, of previously uncultured prokaryotic genomes, and brings greater ownership of all parts of the sequencing process back to individual researchers.

scholarly journals Identification of Pathogenic Structural Variants in Rare Disease Patients through Genome Sequencing

2019 ◽  
Author(s):  
James M. Holt ◽  
Camille L. Birch ◽  
Donna M. Brown ◽  
Manavalan Gajapathy ◽  
Nadiya Sosonkina ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Rare Disease ◽  
Genome Sequencing ◽  
Genomic Analysis ◽  
Whole Genome ◽  
Structural Variants ◽  
Single Nucleotide Variants ◽  
Standard Clinical Practice ◽  
Wide Range ◽  
Genetic Features

AbstractPurposeClinical whole genome sequencing is becoming more common for determining the molecular diagnosis of rare disease. However, standard clinical practice often focuses on small variants such as single nucleotide variants and small insertions/deletions. This leaves a wide range of larger “structural variants” that are not commonly analyzed in patients.MethodsWe developed a pipeline for processing structural variants for patients who received whole genome sequencing through the Undiagnosed Diseases Network (UDN). This pipeline called structural variants, stored them in an internal database, and filtered the variants based on internal frequencies and external annotations. The remaining variants were manually inspected and then interesting findings were reported as research variants to clinical sites in the UDN.ResultsOf 477 analyzed UDN cases, 286 cases (≈ 60%) received at least one structural variant as a research finding. The variants in 16 cases (≈ 4%) are considered “Certain” or “Highly likely” molecularly diagnosed and another 4 cases are currently in review. Of those 20 cases, at least 13 were identified originally through our pipeline with one finding leading to identification of a new disease. As part of this paper, we have also released the collection of variant calls identified in our cohort along with heterozygous and homozygous call counts. This data is available at https://github.com/HudsonAlpha/UDN_SV_export.ConclusionStructural variants are key genetic features that should be analyzed during routine clinical genomic analysis. For our UDN patients, structural variants helped solve ≈ 4% of the total number of cases (≈ 13% of all genome sequencing solves), a success rate we expect to improve with better tools and greater understanding of the human genome.

scholarly journals Application of combined genomic and transfer analyses to identify factors mediating regional spread of antibiotic resistant bacterial lineages

2020 ◽  
Author(s):  
Joyce Wang ◽  
Betsy Foxman ◽  
Ali Pirani ◽  
Zena Lapp ◽  
Lona Mody ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Genomic Analysis ◽  
Patient Characteristics ◽  
Patient Transfer ◽  
Whole Genome ◽  
Nursing Facilities ◽  
Antibiotic Resistant ◽  
Patient Level ◽  
Healthcare Associated

ABSTRACTBackgroundPatients entering nursing facilities (NFs) are frequently colonized with antibiotic resistant organisms (AROs). To understand the determinants of ARO colonization on NF admission we applied whole-genome sequencing to track the spread of four ARO species across regional NFs and evaluated patient-level characteristics and transfer acute-care hospitals (ACHs) as risk factors for colonization.Methods584 patients from six NFs were surveyed for methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus faecalis/faecium (VREfc/VREfm) and ciprofloxacin-resistant Escherichia coli (CipREc) colonization. Genomic analysis was performed to quantify ARO spread between NFs and compared to patient-transfer networks. The association between admission colonization and patient-level variables and recent ACH exposures was examined using multivariable regression models.ResultsThe majority of ARO isolates across study sites belonged to major healthcare-associated lineages: MRSA (ST5;N=89/117); VREfc (ST6;N=68/75); CipREc (ST131; N=58/64), and VREfm (clade A; N=129/129). While the genomic similarity of strains between NF pairs was associated with overlap in their feeder ACHs (Spearman’s rho=0.44-0.75, p<0.05 for MRSA, VREfc and CipREc), limited phylogenetic clustering by either ACH or NF supported regional endemicity. Significant predictors for ARO colonization on NF admission included lower functional status (adjusted odds ratio [aOR]>1 for all four AROs) and recent exposure to glycopeptides (aOR>2 for VREfm, VREfc and MRSA) or 3rd/4th-generation cephalosporins (aOR>2 for MRSA and VREfm). Transfer from specific ACHs was an independent risk factor for only one ARO/ACH pair (VREfm/ACH19, aOR=2.48[1.06-5.83]).ConclusionIn this region, healthcare-associated ARO lineages are endemic among connected NFs and ACHs, making patient characteristics more informative of NF admission colonization risk than exposure to specific ACHs.SummaryUsing a combination of whole-genome sequencing, patient transfer and clinical data, we discerned the dissemination of four high-priority antibiotic-resistant organisms (ARO) in the regional healthcare network, and epidemiolocal drivers underlying the high ARO importation rate into regional nursing facilities.

scholarly journals Identifying transposon insertions in bacterial genomes through nanopore sequencing

2019 ◽  
Author(s):  
David A. Baltrus ◽  
John Medlen ◽  
Meara Clark
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Whole Genome ◽  
Nanopore Sequencing ◽  
Genetic Screens ◽  
Bacterial Genomes ◽  
Sequence Capture ◽  
Transposon Insertion ◽  
Forward Genetic Screens ◽  
Transposon Insertions

AbstractTransposon mutagenesis is a widely used tool for carrying out forward genetic screens across systems, but in some cases it can be difficult to identify transposon insertion points after successful phenotypic screens. As an alternative to traditional methods, we report on the efficacy of using an Oxford Nanopore’s MinION to identify transposon insertions through whole genome sequencing. We also report experiments using CRISPR-Cas to selectively target regions of the genome where a transposon has integrated. Our experiments provide a framework for understanding the efficiency of such techniques for carrying out forward genetic screens and point towards the ability to use CRISPR-based sequence capture to identify the insertion of particular regions of DNA across all genomes, which may enable Tn-Seq experiments using Nanopore based sequencing.

scholarly journals Symptomatic SARS-CoV-2 Reinfection in a Healthy Healthcare Worker in Italy Confirmed by Whole-Genome Sequencing

Viruses ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 899
Author(s):  
Daniela Loconsole ◽  
Anna Sallustio ◽  
Marisa Accogli ◽  
Francesca Centrone ◽  
Daniele Casulli ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Healthcare Worker ◽  
Genome Sequencing ◽  
Genomic Analysis ◽  
Nucleic Acid Amplification ◽  
Whole Genome ◽  
Serological Tests ◽  
Routine Surveillance ◽  
Two Samples ◽  
Genomic Analyses

This study describes a case of SARS-CoV-2 reinfection confirmed by whole-genome sequencing in a healthy physician who had been working in a COVID-19 hospital in Italy since the beginning of the pandemic. Nasopharyngeal swabs were obtained from the patient at each presentation as part of routine surveillance. Nucleic acid amplification testing was performed on the two samples to confirm SARS-CoV-2 infection, and serological tests were used to detect SARS-CoV-2 IgG antibodies. Comparative genome analysis with whole-genome sequencing was performed on nasopharyngeal swabs collected during the two episodes of COVID-19. The first COVID-19 episode was in March 2020, and the second was in January 2021. Both SARS-CoV-2 infections presented with mild symptoms, and seroconversion for SARS-CoV-2 IgG was documented. Genomic analysis showed that the viral genome from the first infection belonged to the lineage B.1.1.74, while that from the second infection to the lineage B.1.177. Epidemiological, clinical, serological, and genomic analyses confirmed that the second episode of SARS-CoV-2 infection in the healthcare worker met the qualifications for “best evidence” for reinfection. Further studies are urgently needed to assess the frequency of such a worrisome occurrence, particularly in the light of the recent diffusion of SARS-CoV-2 variants of concern.

Sign in / Sign up

Export Citation Format

Share Document