An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar

Nathan D. Grubaugh, Karthik Gangavarapu, Joshua Quick, Nathaniel L. Matteson, Jaqueline Goes De Jesus, Bradley Main, Amanda L. Tan, Lauren M. Paul, Doug E. Brackney, Saran Grewal, Nikos Gurfield, Koen K.A. Van Rompay, Sharon Isern, Scott F. Michael, Lark L Schneider, Nicholas J. Loman, Kristian G. Andersen

Research output: Contribution to journalArticle

Abstract

How viruses evolve within hosts can dictate infection outcomes; however, reconstructing this process is challenging. We evaluate our multiplexed amplicon approach, PrimalSeq, to demonstrate how virus concentration, sequencing coverage, primer mismatches, and replicates influence the accuracy of measuring intrahost virus diversity. We develop an experimental protocol and computational tool, iVar, for using PrimalSeq to measure virus diversity using Illumina and compare the results to Oxford Nanopore sequencing. We demonstrate the utility of PrimalSeq by measuring Zika and West Nile virus diversity from varied sample types and show that the accumulation of genetic diversity is influenced by experimental and biological systems.

LanguageEnglish (US)
Article number8
JournalGenome biology
Volume20
Issue number1
DOIs
StatePublished - Jan 8 2019

Fingerprint

virus
Viruses
viruses
Nanopores
West Nile virus
genetic variation
measuring
Infection
infection
sampling

Keywords

  • Amplicon sequencing
  • Intrahost evolution
  • SNP calling
  • Viral sequencing
  • West Nile
  • Zika

ASJC Scopus subject areas

  • Ecology, Evolution, Behavior and Systematics
  • Genetics
  • Cell Biology

Cite this

Grubaugh, N. D., Gangavarapu, K., Quick, J., Matteson, N. L., De Jesus, J. G., Main, B., ... Andersen, K. G. (2019). An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar. Genome biology, 20(1), [8]. https://doi.org/10.1186/s13059-018-1618-7

An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar. / Grubaugh, Nathan D.; Gangavarapu, Karthik; Quick, Joshua; Matteson, Nathaniel L.; De Jesus, Jaqueline Goes; Main, Bradley; Tan, Amanda L.; Paul, Lauren M.; Brackney, Doug E.; Grewal, Saran; Gurfield, Nikos; Van Rompay, Koen K.A.; Isern, Sharon; Michael, Scott F.; Schneider, Lark L; Loman, Nicholas J.; Andersen, Kristian G.

In: Genome biology, Vol. 20, No. 1, 8, 08.01.2019.

Research output: Contribution to journalArticle

Grubaugh, ND, Gangavarapu, K, Quick, J, Matteson, NL, De Jesus, JG, Main, B, Tan, AL, Paul, LM, Brackney, DE, Grewal, S, Gurfield, N, Van Rompay, KKA, Isern, S, Michael, SF, Schneider, LL, Loman, NJ & Andersen, KG 2019, 'An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar', Genome biology, vol. 20, no. 1, 8. https://doi.org/10.1186/s13059-018-1618-7
Grubaugh, Nathan D. ; Gangavarapu, Karthik ; Quick, Joshua ; Matteson, Nathaniel L. ; De Jesus, Jaqueline Goes ; Main, Bradley ; Tan, Amanda L. ; Paul, Lauren M. ; Brackney, Doug E. ; Grewal, Saran ; Gurfield, Nikos ; Van Rompay, Koen K.A. ; Isern, Sharon ; Michael, Scott F. ; Schneider, Lark L ; Loman, Nicholas J. ; Andersen, Kristian G. / An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar. In: Genome biology. 2019 ; Vol. 20, No. 1.
@article{68ea53f63be145029144177ef52baf00,
title = "An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar",
abstract = "How viruses evolve within hosts can dictate infection outcomes; however, reconstructing this process is challenging. We evaluate our multiplexed amplicon approach, PrimalSeq, to demonstrate how virus concentration, sequencing coverage, primer mismatches, and replicates influence the accuracy of measuring intrahost virus diversity. We develop an experimental protocol and computational tool, iVar, for using PrimalSeq to measure virus diversity using Illumina and compare the results to Oxford Nanopore sequencing. We demonstrate the utility of PrimalSeq by measuring Zika and West Nile virus diversity from varied sample types and show that the accumulation of genetic diversity is influenced by experimental and biological systems.",
keywords = "Amplicon sequencing, Intrahost evolution, SNP calling, Viral sequencing, West Nile, Zika",
author = "Grubaugh, {Nathan D.} and Karthik Gangavarapu and Joshua Quick and Matteson, {Nathaniel L.} and {De Jesus}, {Jaqueline Goes} and Bradley Main and Tan, {Amanda L.} and Paul, {Lauren M.} and Brackney, {Doug E.} and Saran Grewal and Nikos Gurfield and {Van Rompay}, {Koen K.A.} and Sharon Isern and Michael, {Scott F.} and Schneider, {Lark L} and Loman, {Nicholas J.} and Andersen, {Kristian G.}",
year = "2019",
month = "1",
day = "8",
doi = "10.1186/s13059-018-1618-7",
language = "English (US)",
volume = "20",
journal = "Genome Biology",
issn = "1465-6914",
publisher = "BioMed Central",
number = "1",

}

TY - JOUR

T1 - An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar

AU - Grubaugh, Nathan D.

AU - Gangavarapu, Karthik

AU - Quick, Joshua

AU - Matteson, Nathaniel L.

AU - De Jesus, Jaqueline Goes

AU - Main, Bradley

AU - Tan, Amanda L.

AU - Paul, Lauren M.

AU - Brackney, Doug E.

AU - Grewal, Saran

AU - Gurfield, Nikos

AU - Van Rompay, Koen K.A.

AU - Isern, Sharon

AU - Michael, Scott F.

AU - Schneider, Lark L

AU - Loman, Nicholas J.

AU - Andersen, Kristian G.

PY - 2019/1/8

Y1 - 2019/1/8

N2 - How viruses evolve within hosts can dictate infection outcomes; however, reconstructing this process is challenging. We evaluate our multiplexed amplicon approach, PrimalSeq, to demonstrate how virus concentration, sequencing coverage, primer mismatches, and replicates influence the accuracy of measuring intrahost virus diversity. We develop an experimental protocol and computational tool, iVar, for using PrimalSeq to measure virus diversity using Illumina and compare the results to Oxford Nanopore sequencing. We demonstrate the utility of PrimalSeq by measuring Zika and West Nile virus diversity from varied sample types and show that the accumulation of genetic diversity is influenced by experimental and biological systems.

AB - How viruses evolve within hosts can dictate infection outcomes; however, reconstructing this process is challenging. We evaluate our multiplexed amplicon approach, PrimalSeq, to demonstrate how virus concentration, sequencing coverage, primer mismatches, and replicates influence the accuracy of measuring intrahost virus diversity. We develop an experimental protocol and computational tool, iVar, for using PrimalSeq to measure virus diversity using Illumina and compare the results to Oxford Nanopore sequencing. We demonstrate the utility of PrimalSeq by measuring Zika and West Nile virus diversity from varied sample types and show that the accumulation of genetic diversity is influenced by experimental and biological systems.

KW - Amplicon sequencing

KW - Intrahost evolution

KW - SNP calling

KW - Viral sequencing

KW - West Nile

KW - Zika

UR - http://www.scopus.com/inward/record.url?scp=85059769331&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85059769331&partnerID=8YFLogxK

U2 - 10.1186/s13059-018-1618-7

DO - 10.1186/s13059-018-1618-7

M3 - Article

VL - 20

JO - Genome Biology

T2 - Genome Biology

JF - Genome Biology

SN - 1465-6914

IS - 1

M1 - 8

ER -