scholarly journals Contamination-controlled high-throughput whole genome sequencing for influenza A viruses using the MiSeq sequencer

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Hong Kai Lee ◽  
Chun Kiat Lee ◽  
Julian Wei-Tze Tang ◽  
Tze Ping Loh ◽  
Evelyn Siew-Chuan Koay
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
High Throughput ◽  
Influenza A ◽  
Whole Genome ◽  
Influenza A Viruses

scholarly journals Efficient whole genome sequencing of influenza A viruses

2019 ◽  
Author(s):  
Marina Escalera-Zamudio ◽  
Ana Georgina Cobián-Güemes ◽  
Blanca Taboada ◽  
Irma López-Martínez ◽  
Joel Armando Vázquez-Pérez ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Influenza A Virus ◽  
Genome Sequencing ◽  
Influenza A ◽  
Genetic Material ◽  
Influenza Viruses ◽  
Epidemiological Surveillance ◽  
Clinical Samples ◽  
Whole Genome ◽  
Influenza A Viruses

ABSTRACTThe constant threat of emergence for novel pathogenic influenza A viruses with pandemic potential, makes full-genome characterization of circulating influenza viral strains a high priority, allowing detection of novel and re-assorting variants. Sequencing the full-length genome of influenza A virus traditionally required multiple amplification rounds, followed by the subsequent sequencing of individual PCR products. The introduction of high-throughput sequencing technologies has made whole genome sequencing easier and faster. We present a simple protocol to obtain whole genome sequences of hypothetically any influenza A virus, even with low quantities of starting genetic material. The complete genomes of influenza A viruses of different subtypes and from distinct sources (clinical samples of pdmH1N1, tissue culture-adapted H3N2 viruses, or avian influenza viruses from cloacal swabs) were amplified with a single multisegment reverse transcription-PCR reaction and sequenced using Illumina sequencing platform. Samples with low quantity of genetic material after initial PCR amplification were re-amplified by an additional PCR using random primers. Whole genome sequencing was successful for 66% of the samples, whilst the most relevant genome segments for epidemiological surveillance (corresponding to the hemagglutinin and neuraminidase) were sequenced with at least 93% coverage (and a minimum 10x) for 98% of the samples. Low coverage for some samples is likely due to an initial low viral RNA concentration in the original sample. The proposed methodology is especially suitable for sequencing a large number of samples, when genetic data is urgently required for strains characterization, and may also be useful for variant analysis.

scholarly journals Surveillance of Influenza A resistance to polymerase complex inhibitors by whole genome analysis, 2016-17, 2017-18 and 2018-19 Seasons, Eastern Spain

2021 ◽  
Author(s):  
Beatriz Mengual-Chuliá ◽  
Andrés Alonso-Cordero ◽  
Laura Cano ◽  
M. del Mar Mosquera ◽  
Patricia de Molina ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Genome Analysis ◽  
Influenza A ◽  
Natural Resistance ◽  
Whole Genome ◽  
Influenza A Viruses ◽  
Whole Genome Analysis ◽  
Molecular Surveillance ◽  
Polymerase Complex

Molecular surveillance by whole genome sequencing was used to monitor the susceptibility of circulating Influenza A viruses to three polymerase complex inhibitors. A total of 12 resistance substitutions were found among 285 genomes analysed, but none associated with high levels of resistance. Natural resistance to these influenza A antivirals is currently uncommon.

scholarly journals Method comparison of targeted influenza A virus typing and whole-genome sequencing from respiratory specimens of companion animals

2020 ◽  
pp. 104063872093387
Author(s):  
Patrick K. Mitchell ◽  
Brittany D. Cronk ◽  
Ian E. H. Voorhees ◽  
Derek Rothenheber ◽  
Renee R. Anderson ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Influenza A ◽  
Sequence Data ◽  
Companion Animals ◽  
The United States ◽  
Quality Data ◽  
Whole Genome ◽  
Influenza A Viruses ◽  
Respiratory Specimens

Epidemics of H3N8 and H3N2 influenza A viruses (IAVs) in dogs, along with recognition of spillover infections from IAV strains typically found in humans or other animals, have emphasized the importance of efficient laboratory testing. Given the lack of active IAV surveillance or immunization requirements for dogs, cats, or horses imported into the United States, serotype prediction and whole-genome sequencing of positive specimens detected at veterinary diagnostic laboratories are also needed. The conserved sequences at the ends of the viral genome segments facilitate universal amplification of all segments of viral genomes directly from respiratory specimens. Although several methods for genomic analysis have been reported, no optimization focusing on companion animal strains has been described, to our knowledge. We compared 2 sets of published universal amplification primers using 26 IAV-positive specimens from dogs, horses, and a cat. Libraries prepared from the resulting amplicons were sequenced using Illumina chemistry, and reference-based assemblies were generated from the data produced by both methods. Although both methods produced high-quality data, coverage profiles and base calling differed between the 2 methods. The sequence data were also used to identify the subtype of the IAV strains sequenced and then compared to standard PCR assays for neuraminidase types N2 and N8.

scholarly journals Evaluation of two workflows for whole genome sequencing-based typing of influenza A viruses

2019 ◽  
Vol 266 ◽  
pp. 30-33 ◽  
Author(s):  
Daniel Wüthrich ◽  
Daniela Lang ◽  
Nicola F. Müller ◽  
Richard A. Neher ◽  
Tanja Stadler ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Influenza A ◽  
Whole Genome ◽  
Influenza A Viruses

Rapid, high throughput library preparation of SARS-CoV2 using Illumina’s DNA Prep Library Preparation Kit v2

2022 ◽  
Author(s):  
Jason Nguyen ◽  
Rebecca Hickman ◽  
Tracy Lee ◽  
Natalie Prystajecky ◽  
John Tyson
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
High Throughput ◽  
Illumina Miseq ◽  
Whole Genome ◽  
Library Preparation ◽  
Dna Libraries ◽  
Reaction Volumes ◽  
Single Size ◽  
Stop Buffer

This procedure provides instructions on how to prepare DNA libraries for whole genome sequencing on an Illumina MiSeq or NextSeq using Illumina’s DNA Prep Library Preparation Kit scaled to half reaction volumes with modifications to the post-PCR procedures; tagmentation stop buffer and associated washes are removed and libraries are pooled post PCR then a single size selection is performed. This protocol is used to sequence SARS-CoV-2 using the cDNA/PCR protocol: https://dx.doi.org/10.17504/protocols.io.b3viqn4e

scholarly journals High-throughput screening and whole genome sequencing identifies an antimicrobially active inhibitor of Vibrio cholerae

2014 ◽  
Vol 14 (1) ◽  
pp. 49 ◽  
Author(s):  
Galina Sergeev ◽  
Sambit Roy ◽  
Michael Jarek ◽  
Viktor Zapolskii ◽  
Dieter E Kaufmann ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Vibrio Cholerae ◽  
Genome Sequencing ◽  
High Throughput ◽  
High Throughput Screening ◽  
Whole Genome ◽  
Active Inhibitor

scholarly journals Integrating patient and whole genome sequencing data to provide insights into the epidemiology of seasonal influenza A(H3N2) viruses

2017 ◽  
Author(s):  
Emily J. Goldstein ◽  
William T. Harvey ◽  
Gavin S. Wilkie ◽  
Samantha J. Shepherd ◽  
Alasdair R. MacLean ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Influenza A ◽  
Seasonal Influenza ◽  
Epidemiological Data ◽  
Respiratory Illness ◽  
Influenza Surveillance ◽  
Whole Genome Sequencing Data ◽  
Whole Genome ◽  
Sequencing Data

AbstractGenetic surveillance of seasonal influenza is largely focused upon sequencing of the haemagglutinin gene. Consequently, our understanding of the contribution of the remaining seven gene segments to the evolution and epidemiological dynamics of seasonal influenza is relatively limited. The increased availability of next generation sequencing technologies allows rapid and economic whole genome sequencing (WGS). Here, 150 influenza A(H3N2) positive clinical specimens with linked epidemiological data, from the 2014/15 season in Scotland, were sequenced directly using both Sanger sequencing of the HA1 region and WGS using the Illumina MiSeq platform. Sequences generated by both methods were highly consistent and WGS provided on average >90% whole genome coverage. As reported in other European countries during 2014/15, all strains belonged to genetic group 3C, with subgroup 3C.2a predominating. Inter-subgroup reassortants were identified (9%), including three 3C.3 viruses descended from a single reassortment event, which had persisted in the population. Significant phylogenetic associations with cases of severe acute respiratory illness observed herein warrant further investigation. Severe cases were also more likely to be associated with reassortant viruses (odds ratio: 4.4 (1.3-15.5)) and occur later in the season. These results suggest that increased levels of WGS, linked to clinical and epidemiological data, could improve influenza surveillance.

Sign in / Sign up

Export Citation Format

Share Document