Nanopore sequencing from extraction-free direct PCR of dried serum spots for portable hepatitis B virus drug-resistance typing

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Autor(es): dc.contributorUniv Nottingham-
Autor(es): dc.contributorNottingham Univ Hosp NHS Trust-
Autor(es): dc.contributorBrazilian Minist Hlth-
Autor(es): dc.contributorUniversidade Federal de Uberlândia (UFU)-
Autor(es): dc.contributorUniv Sulaimani-
Autor(es): dc.contributorHawler Med Univ-
Autor(es): dc.contributorCent Publ Hlth Lab-
Autor(es): dc.contributorUniversidade Estadual Paulista (Unesp)-
Autor(es): dc.creatorAstbury, Stuart-
Autor(es): dc.creatorNunes Soares, Marcia Maria Costa-
Autor(es): dc.creatorPeprah, Emmanuel-
Autor(es): dc.creatorKing, Barnabas-
Autor(es): dc.creatorGomes Jardim, Ana Carolina [UNESP]-
Autor(es): dc.creatorFarinha Shimizu, Jacqueline [UNESP]-
Autor(es): dc.creatorJalal, Paywast-
Autor(es): dc.creatorSaeed, Chiman H.-
Autor(es): dc.creatorSabeer, Furat T.-
Autor(es): dc.creatorIrving, William L.-
Autor(es): dc.creatorTarr, Alexander W.-
Autor(es): dc.creatorMcClure, C. Patrick-
Data de aceite: dc.date.accessioned2022-02-22T00:06:08Z-
Data de disponibilização: dc.date.available2022-02-22T00:06:08Z-
Data de envio: dc.date.issued2020-12-09-
Data de envio: dc.date.issued2020-12-09-
Data de envio: dc.date.issued2020-08-01-
Fonte completa do material: dc.identifierhttp://dx.doi.org/10.1016/j.jcv.2020.104483-
Fonte completa do material: dc.identifierhttp://hdl.handle.net/11449/195547-
Fonte: dc.identifier.urihttp://educapes.capes.gov.br/handle/11449/195547-
Descrição: dc.descriptionBackground: Effective drug regimens for the treatment of hepatitis B virus (HBV) infections are essential to achieve the World Health Organisation commitment to eliminate viral hepatitis by 2030. Lamivudine (3TC) is widely used in countries with high levels of chronic HBV, however resistance has been shown to occur in up to 50 % of individuals receiving continuous monotherapy for 4 years. Telbivudine (LdT) is now more commonly used in place of lamivudine but is ineffective against 3TC-resistant HBV. Genotyping and identification of resistance-associated substitutions (RAS) is not practical in many locations. Objectives: A novel assay was designed to enable HBV genotyping and characterisation of resistance mutations directly from serum samples stored on filter paper, using Sanger and MinION sequencing. Study design: The assay was applied to a cohort of 30 samples stored on filter paper for several years with HBV viral loads ranging from 8.2 x 10(8) to 635 IU/mL. A set of 6 high-titre samples were used in a proof-of-principle study using the MinION sequencer. Results: The assay allowed determination of HBV genotype and elucidation of RAS down to 600 IU/mL using a 550bp amplicon. Sequencing of a 1.2 kb amplicon using a MinION sequencer gave results consistent with Sanger sequencing and allowed the identification of minor populations of variants. Conclusions: We present two approaches for reliable HBV sequencing and RAS identification using methods suitable for resource-limited environments. This is the first demonstration of extraction-free DNA sequencing direct from DSS using MinION and these workflows are adaptable to the investigation of other DNA viruses.-
Descrição: dc.descriptionNottingham Molecular Pathology Node (United Kingdom MRC/EPSRC)-
Descrição: dc.descriptionNational Institute for Health Research (NIHR) Nottingham Biomedical Research Centre-
Descrição: dc.descriptionUniv Nottingham, Nottingham Digest Dis Ctr, Sch Med, Nottingham, England-
Descrição: dc.descriptionNottingham Univ Hosp NHS Trust, NIHR Nottingham Biomed Res Ctr, Nottingham, England-
Descrição: dc.descriptionUniv Nottingham, Nottingham, England-
Descrição: dc.descriptionUniv Nottingham, MRC EPSRC Nottingham Mol Pathol Node, Nottingham, England-
Descrição: dc.descriptionBrazilian Minist Hlth, Inst Adolfo Lutz, Sao Paulo, SP, Brazil-
Descrição: dc.descriptionUniv Nottingham, Sch Life Sci, Nottingham, England-
Descrição: dc.descriptionUniv Fed Uberlandia, Inst Biomed Sci, Uberlandia, MG, Brazil-
Descrição: dc.descriptionUniv Sulaimani, Fac Sci, Biol Dept, Sulaymaniyah, Iraq-
Descrição: dc.descriptionHawler Med Univ, Med Res Ctr, Erbil, Iraq-
Descrição: dc.descriptionCent Publ Hlth Lab, Erbil, Iraq-
Descrição: dc.descriptionSao Paulo State Univ, IBILCE, Sao Jose Do Rio Preto, SP, Brazil-
Descrição: dc.descriptionSao Paulo State Univ, IBILCE, Sao Jose Do Rio Preto, SP, Brazil-
Descrição: dc.descriptionNottingham Molecular Pathology Node (United Kingdom MRC/EPSRC): MR/N005953/1-
Formato: dc.format7-
Idioma: dc.languageen-
Publicador: dc.publisherElsevier B.V.-
Relação: dc.relationJournal Of Clinical Virology-
???dc.source???: dc.sourceWeb of Science-
Palavras-chave: dc.subjectDried blood spot-
Palavras-chave: dc.subjectNanopore sequencing-
Palavras-chave: dc.subjectPortable diagnostic test-
Palavras-chave: dc.subjectHepatitis B virus-
Palavras-chave: dc.subjectResistance-associated substitutions-
Palavras-chave: dc.subjectGenotyping-
Título: dc.titleNanopore sequencing from extraction-free direct PCR of dried serum spots for portable hepatitis B virus drug-resistance typing-
Tipo de arquivo: dc.typelivro digital-
Aparece nas coleções:Repositório Institucional - Unesp

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