Endless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts

ABSTRACT RNA viruses are infamous for their high rates of mutation, which produce swarms of genetic variants within individual hosts. To date, analyses of intrahost genetic diversity have focused on the primary genome sequence. However, virus phenotypes are shaped not only by primary sequence but al...

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Autores principales:	Stacey L. P. Scroggs, Nathan D. Grubaugh, Johnny A. Sena, Anitha Sundararajan, Faye D. Schilkey, Darci R. Smith, Gregory D. Ebel, Kathryn A. Hanley
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	RNA virus West Nile virus evolution genetic diversity natural selection quasispecies Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/ce12721eeb934822a1fd589614d2b0e1
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spelling	oai:doaj.org-article:ce12721eeb934822a1fd589614d2b0e12021-11-15T15:22:20ZEndless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts10.1128/mSphere.00291-192379-5042https://doaj.org/article/ce12721eeb934822a1fd589614d2b0e12019-06-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mSphere.00291-19https://doaj.org/toc/2379-5042ABSTRACT RNA viruses are infamous for their high rates of mutation, which produce swarms of genetic variants within individual hosts. To date, analyses of intrahost genetic diversity have focused on the primary genome sequence. However, virus phenotypes are shaped not only by primary sequence but also by the secondary structures into which this sequence folds. Such structures enable viral replication, translation, and binding of small RNAs, yet within-host variation at the structural level has not been adequately explored. We characterized the structural diversity of the 5′ untranslated region (UTR) of populations of West Nile virus (WNV) that had been subject to five serial passages in triplicate in each of three bird species. Viral genomes were sampled from host serum samples at each passage (n = 45 populations) and subjected to next-generation sequencing. For populations derived from passages 1, 3, and 5 (n = 9 populations), we predicted the impact of each mutation occurring at a frequency of ≥1% on the secondary structure of the 5′ UTR. As expected, mutations in double-stranded (DS) regions of the 5′ UTR stem structures caused structural changes of significantly greater magnitude than did mutations in single-stranded (SS) regions. Despite the greater impact of mutations in DS regions, mutations in DS and SS regions occurred at similar frequencies, with no evidence of enhanced selection against mutation in DS regions. In contrast, mutations in two regions that mediate genome cyclization and thereby regulate replication and translation, the 5′ cyclization sequence and the UAR flanking stem (UFS), were suppressed in all three hosts. IMPORTANCE The enzymes that copy RNA genomes lack proofreading, and viruses that possess RNA genomes, such as West Nile virus, rapidly diversify into swarms of mutant lineages within a host. Intrahost variation of the primary genomic sequence of RNA viruses has been studied extensively because the extent of this variation shapes key virus phenotypes. However, RNA genomes also form complex secondary structures based on within-genome nucleotide complementarity, which are critical regulators of the cyclization of the virus genome that is necessary for efficient replication and translation. We sought to characterize variation in these secondary structures within populations of West Nile virus during serial passage in three bird species. Our study indicates that the intrahost population of West Nile virus is a diverse assortment of RNA secondary structures that should be considered in future analyses of intrahost viral diversity, but some regions that are critical for genome cyclization are conserved within hosts. Besides potential impacts on viral replication, structural diversity can influence the efficacy of small RNA antiviral therapies.Stacey L. P. ScroggsNathan D. GrubaughJohnny A. SenaAnitha SundararajanFaye D. SchilkeyDarci R. SmithGregory D. EbelKathryn A. HanleyAmerican Society for MicrobiologyarticleRNA virusWest Nile virusevolutiongenetic diversitynatural selectionquasispeciesMicrobiologyQR1-502ENmSphere, Vol 4, Iss 3 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	RNA virus West Nile virus evolution genetic diversity natural selection quasispecies Microbiology QR1-502
spellingShingle	RNA virus West Nile virus evolution genetic diversity natural selection quasispecies Microbiology QR1-502 Stacey L. P. Scroggs Nathan D. Grubaugh Johnny A. Sena Anitha Sundararajan Faye D. Schilkey Darci R. Smith Gregory D. Ebel Kathryn A. Hanley Endless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts
description	ABSTRACT RNA viruses are infamous for their high rates of mutation, which produce swarms of genetic variants within individual hosts. To date, analyses of intrahost genetic diversity have focused on the primary genome sequence. However, virus phenotypes are shaped not only by primary sequence but also by the secondary structures into which this sequence folds. Such structures enable viral replication, translation, and binding of small RNAs, yet within-host variation at the structural level has not been adequately explored. We characterized the structural diversity of the 5′ untranslated region (UTR) of populations of West Nile virus (WNV) that had been subject to five serial passages in triplicate in each of three bird species. Viral genomes were sampled from host serum samples at each passage (n = 45 populations) and subjected to next-generation sequencing. For populations derived from passages 1, 3, and 5 (n = 9 populations), we predicted the impact of each mutation occurring at a frequency of ≥1% on the secondary structure of the 5′ UTR. As expected, mutations in double-stranded (DS) regions of the 5′ UTR stem structures caused structural changes of significantly greater magnitude than did mutations in single-stranded (SS) regions. Despite the greater impact of mutations in DS regions, mutations in DS and SS regions occurred at similar frequencies, with no evidence of enhanced selection against mutation in DS regions. In contrast, mutations in two regions that mediate genome cyclization and thereby regulate replication and translation, the 5′ cyclization sequence and the UAR flanking stem (UFS), were suppressed in all three hosts. IMPORTANCE The enzymes that copy RNA genomes lack proofreading, and viruses that possess RNA genomes, such as West Nile virus, rapidly diversify into swarms of mutant lineages within a host. Intrahost variation of the primary genomic sequence of RNA viruses has been studied extensively because the extent of this variation shapes key virus phenotypes. However, RNA genomes also form complex secondary structures based on within-genome nucleotide complementarity, which are critical regulators of the cyclization of the virus genome that is necessary for efficient replication and translation. We sought to characterize variation in these secondary structures within populations of West Nile virus during serial passage in three bird species. Our study indicates that the intrahost population of West Nile virus is a diverse assortment of RNA secondary structures that should be considered in future analyses of intrahost viral diversity, but some regions that are critical for genome cyclization are conserved within hosts. Besides potential impacts on viral replication, structural diversity can influence the efficacy of small RNA antiviral therapies.
format	article
author	Stacey L. P. Scroggs Nathan D. Grubaugh Johnny A. Sena Anitha Sundararajan Faye D. Schilkey Darci R. Smith Gregory D. Ebel Kathryn A. Hanley
author_facet	Stacey L. P. Scroggs Nathan D. Grubaugh Johnny A. Sena Anitha Sundararajan Faye D. Schilkey Darci R. Smith Gregory D. Ebel Kathryn A. Hanley
author_sort	Stacey L. P. Scroggs
title	Endless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts
title_short	Endless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts
title_full	Endless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts
title_fullStr	Endless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts
title_full_unstemmed	Endless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts
title_sort	endless forms: within-host variation in the structure of the west nile virus rna genome during serial passage in bird hosts
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/ce12721eeb934822a1fd589614d2b0e1
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Endless Forms: Within-Host Variation in the Structure of the West Nile Virus RNA Genome during Serial Passage in Bird Hosts

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