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Development of an amplicon-based sequencing approach in response to the global emergence of mpox

2023-06-13Journal article Open access
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dc.contributor.authorChen, Nicholas F.G.
dc.contributor.authorChaguza, Chrispin
dc.contributor.authorGagne, Luc
dc.contributor.authorDoucette, Matthew
dc.contributor.authorSmole, Sandra
dc.contributor.authorBuzby, Erika
dc.contributor.authorHall, Joshua
dc.contributor.authorAsh, Stephanie
dc.contributor.authorHarrington, Rachel
dc.contributor.authorCofsky, Seana
dc.contributor.authorClancy, Selina
dc.contributor.authorKapsak, Curtis J.
dc.contributor.authorSevinsky, Joel
dc.contributor.authorLibuit, Kevin
dc.contributor.authorPark, Daniel J.
dc.contributor.authorHemarajata, Peera
dc.contributor.authorGarrigues, Jacob M.
dc.contributor.authorGreen, Nicole M.
dc.contributor.authorSierra-Patev, Sean
dc.contributor.authorCarpenter-Azevedo, Kristin
dc.contributor.authorHuard, Richard C.
dc.contributor.authorPearson, Claire
dc.contributor.authorIncekara, Kutluhan
dc.contributor.authorNishimura, Christina
dc.contributor.authorHuang, Jian Ping
dc.contributor.authorGagnon, Emily
dc.contributor.authorReever, Ethan
dc.contributor.authorRazeq, Jafar
dc.contributor.authorMuyombwe, Anthony
dc.contributor.authorBorges, Vítor
dc.contributor.authorFerreira, Rita
dc.contributor.authorSobral, Daniel
dc.contributor.authorDuarte, Silvia
dc.contributor.authorSantos, Daniela
dc.contributor.authorVieira, Luís
dc.contributor.authorGomes, João Paulo
dc.contributor.authorAquino, Carly
dc.contributor.authorSavino, Isabella M.
dc.contributor.authorFelton, Karinda
dc.contributor.authorBajwa, Moneeb
dc.contributor.authorHayward, Nyjil
dc.contributor.authorMiller, Holly
dc.contributor.authorNaumann, Allison
dc.contributor.authorAllman, Ria
dc.contributor.authorGreer, Neel
dc.contributor.authorFall, Amary
dc.contributor.authorMostafa, Heba H.
dc.contributor.authorMcHugh, Martin P.
dc.contributor.authorMaloney, Daniel M.
dc.contributor.authorDewar, Rebecca
dc.contributor.authorKenicer, Juliet
dc.contributor.authorParker, Abby
dc.contributor.authorMathers, Katharine
dc.contributor.authorWild, Jonathan
dc.contributor.authorCotton, Seb
dc.contributor.authorTempleton, Kate E.
dc.contributor.authorChurchwell, George
dc.contributor.authorLee, Philip A.
dc.contributor.authorPedrosa, Maria
dc.contributor.authorMcGruder, Brenna
dc.contributor.authorSchmedes, Sarah
dc.contributor.authorPlumb, Matthew R.
dc.contributor.authorWang, Xiong
dc.contributor.authorBarcellos, Regina Bones
dc.contributor.authorGodinho, Fernanda M.S.
dc.contributor.authorSalvato, Richard Steiner
dc.contributor.authorCeniseros, Aimee
dc.contributor.authorBreban, Mallery I.
dc.contributor.authorGrubaugh, Nathan D.
dc.contributor.authorGallagher, Glen R.
dc.contributor.authorVogels, Chantal B.F.
dc.date.accessioned2024-01-23T11:54:36Z
dc.date.available2024-01-23T11:54:36Z
dc.date.issued2023-06-13
dc.description.abstractThe 2022 multicountry mpox outbreak concurrent with the ongoing Coronavirus Disease 2019 (COVID-19) pandemic further highlighted the need for genomic surveillance and rapid pathogen whole-genome sequencing. While metagenomic sequencing approaches have been used to sequence many of the early mpox infections, these methods are resource intensive and require samples with high viral DNA concentrations. Given the atypical clinical presentation of cases associated with the outbreak and uncertainty regarding viral load across both the course of infection and anatomical body sites, there was an urgent need for a more sensitive and broadly applicable sequencing approach. Highly multiplexed amplicon-based sequencing (PrimalSeq) was initially developed for sequencing of Zika virus, and later adapted as the main sequencing approach for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). Here, we used PrimalScheme to develop a primer scheme for human monkeypox virus that can be used with many sequencing and bioinformatics pipelines implemented in public health laboratories during the COVID-19 pandemic. We sequenced clinical specimens that tested presumptively positive for human monkeypox virus with amplicon-based and metagenomic sequencing approaches. We found notably higher genome coverage across the virus genome, with minimal amplicon drop-outs, in using the amplicon-based sequencing approach, particularly in higher PCR cycle threshold (Ct) (lower DNA titer) samples. Further testing demonstrated that Ct value correlated with the number of sequencing reads and influenced the percent genome coverage. To maximize genome coverage when resources are limited, we recommend selecting samples with a PCR Ct below 31 Ct and generating 1 million sequencing reads per sample. To support national and international public health genomic surveillance efforts, we sent out primer pool aliquots to 10 laboratories across the United States, United Kingdom, Brazil, and Portugal. These public health laboratories successfully implemented the human monkeypox virus primer scheme in various amplicon sequencing workflows and with different sample types across a range of Ct values. Thus, we show that amplicon-based sequencing can provide a rapidly deployable, cost-effective, and flexible approach to pathogen whole-genome sequencing in response to newly emerging pathogens. Importantly, through the implementation of our primer scheme into existing SARS-CoV-2 workflows and across a range of sample types and sequencing platforms, we further demonstrate the potential of this approach for rapid outbreak response.pt_PT
dc.description.sponsorshipThis publication was made possible by CTSA Grant Number UL1 TR001863 from the National Center for Advancing Translational Science (NCATS), a component of the National Institutes of Health (NIH) awarded to CBFV. INSA was partially funded by the HERA project (Grant/ 2021/PHF/23776) supported by the European Commission through the European Centre for Disease Control (to VB).pt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.citationPLoS Biol. 2023 Jun 13;21(6):e3002151. doi: 10.1371/journal.pbio.3002151. eCollection 2023 Junpt_PT
dc.identifier.doi10.1371/journal.pbio.3002151pt_PT
dc.identifier.issn1544-9173
dc.identifier.urihttp://hdl.handle.net/10400.18/8955
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherPublic Library of Sciencept_PT
dc.relation.publisherversionhttps://journals.plos.org/plosbiology/article?id=10.1371/journal.pbio.3002151pt_PT
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/pt_PT
dc.subjectMpoxpt_PT
dc.subjectMpox Outbreakpt_PT
dc.subjectMonkeypox Viruspt_PT
dc.subjectGenomic Surveillancept_PT
dc.subjectWhole-genome Sequencingpt_PT
dc.subjectOutbreak Responsept_PT
dc.subjectGlobal Emergencept_PT
dc.subjectPandemicspt_PT
dc.subjectInfecções Emergentespt_PT
dc.titleDevelopment of an amplicon-based sequencing approach in response to the global emergence of mpoxpt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.citation.issue6pt_PT
oaire.citation.startPagee3002151pt_PT
oaire.citation.titlePLOS Biologypt_PT
oaire.citation.volume21pt_PT
rcaap.embargofctAcesso de acordo com a política editorial da revista.pt_PT
rcaap.rightsopenAccesspt_PT
rcaap.typearticlept_PT

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