Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus

Autores
Tomás, Gonzalo; Hernández, Martín; Marandino, Ana; Panzera, Yanina; Maya, Leticia; Hernández, Diego; Pereda, Ariel Julián; Banda, Alejandro; Villegas, Pedro; Aguirre, Sebastian; Pérez, Ruben
Año de publicación
2012
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Rapid and reliable detection and classification of infectious bursal disease viruses (IBDVs) is of crucial importance for disease surveillance and control. This study presents the development and validation of a real-time RT-PCR assay to detect and discriminate very virulent (vv) from non-vv (classic and variant) IBDV strains. The assay uses two fluorogenic, minor groove-binding (MGB) TaqMan probes targeted to a single nucleotide polymorphism (SNP) embedded in a highly conserved genomic region. The analyti- cal sensitivity of the assay was determined using serial dilutions of in vitro-transcribed RNA. The assay demonstrated a wide dynamic range between 102 and 108 standard RNA copies per reaction. Good repro- ducibility was also detected, with intra- and inter-assay coefficients of variation ranging from 0.13% to 2.23% and 0.26% to 1.92%, respectively. The assay detected successfully all the assessed vv, classical, and variant field and vaccine strains and correctly discriminated all vvIBDV strains from non-vvIBDV strains. Other common avian RNA viruses tested negative, indicating high specificity of the assay. The high sensi- tivity, rapidity, reproducibility, and specificity of the real-time RT-PCR assay make this method suitable for general and genotype-specific detection and quantitation.
Fil: Tomás, Gonzalo. Universidad del Uruguay. Facultad de Ciencias; Uruguay
Fil: Hernández, Martín. Universidad del Uruguay. Facultad de Ciencias; Uruguay
Fil: Marandino, Ana. Universidad del Uruguay. Facultad de Ciencias; Uruguay
Fil: Panzera, Yanina. Universidad de la República. Facultad de Ciencias; Uruguay
Fil: Maya, Leticia. Universidad de la República. Facultad de Ciencias; Uruguay
Fil: Hernández, Diego. Universidad de la República. Facultad de Ciencias; Uruguay
Fil: Pereda, Ariel Julián. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Banda, Alejandro. Mississippi State University; Estados Unidos
Fil: Villegas, Pedro. Georgia State University; Estados Unidos
Fil: Aguirre, Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencia y Tecnología "Dr. César Milstein". Fundación Pablo Cassará. Instituto de Ciencia y Tecnología "Dr. César Milstein"; Argentina
Fil: Pérez, Ruben. Universidad de la República. Facultad de Ciencias; Uruguay
Materia
DIAGNOSIS
IBDV
INFECTIOUS BURSAL DISEASE (GUMBORO)
REAL-TIME PCR
STRAIN CHARACTERIZATION
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/197963
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/197963
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virusTomás, GonzaloHernández, MartínMarandino, AnaPanzera, YaninaMaya, LeticiaHernández, DiegoPereda, Ariel JuliánBanda, AlejandroVillegas, PedroAguirre, SebastianPérez, RubenDIAGNOSISIBDVINFECTIOUS BURSAL DISEASE (GUMBORO)REAL-TIME PCRSTRAIN CHARACTERIZATIONhttps://purl.org/becyt/ford/4.3https://purl.org/becyt/ford/4Rapid and reliable detection and classification of infectious bursal disease viruses (IBDVs) is of crucial importance for disease surveillance and control. This study presents the development and validation of a real-time RT-PCR assay to detect and discriminate very virulent (vv) from non-vv (classic and variant) IBDV strains. The assay uses two fluorogenic, minor groove-binding (MGB) TaqMan probes targeted to a single nucleotide polymorphism (SNP) embedded in a highly conserved genomic region. The analyti- cal sensitivity of the assay was determined using serial dilutions of in vitro-transcribed RNA. The assay demonstrated a wide dynamic range between 102 and 108 standard RNA copies per reaction. Good repro- ducibility was also detected, with intra- and inter-assay coefficients of variation ranging from 0.13% to 2.23% and 0.26% to 1.92%, respectively. The assay detected successfully all the assessed vv, classical, and variant field and vaccine strains and correctly discriminated all vvIBDV strains from non-vvIBDV strains. Other common avian RNA viruses tested negative, indicating high specificity of the assay. The high sensi- tivity, rapidity, reproducibility, and specificity of the real-time RT-PCR assay make this method suitable for general and genotype-specific detection and quantitation.Fil: Tomás, Gonzalo. Universidad del Uruguay. Facultad de Ciencias; UruguayFil: Hernández, Martín. Universidad del Uruguay. Facultad de Ciencias; UruguayFil: Marandino, Ana. Universidad del Uruguay. Facultad de Ciencias; UruguayFil: Panzera, Yanina. Universidad de la República. Facultad de Ciencias; UruguayFil: Maya, Leticia. Universidad de la República. Facultad de Ciencias; UruguayFil: Hernández, Diego. Universidad de la República. Facultad de Ciencias; UruguayFil: Pereda, Ariel Julián. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Banda, Alejandro. Mississippi State University; Estados UnidosFil: Villegas, Pedro. Georgia State University; Estados UnidosFil: Aguirre, Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencia y Tecnología "Dr. César Milstein". Fundación Pablo Cassará. Instituto de Ciencia y Tecnología "Dr. César Milstein"; ArgentinaFil: Pérez, Ruben. Universidad de la República. Facultad de Ciencias; UruguayElsevier Science2012-10info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/197963Tomás, Gonzalo; Hernández, Martín; Marandino, Ana; Panzera, Yanina; Maya, Leticia; et al.; Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus; Elsevier Science; Journal of Virological Methods; 185; 1; 10-2012; 101-1070166-0934CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0166093412002108info:eu-repo/semantics/altIdentifier/doi/10.1016/j.jviromet.2012.06.012info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-03T09:58:02Zoai:ri.conicet.gov.ar:11336/197963instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-03 09:58:02.309CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus
title Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus
spellingShingle Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus
Tomás, Gonzalo
DIAGNOSIS
IBDV
INFECTIOUS BURSAL DISEASE (GUMBORO)
REAL-TIME PCR
STRAIN CHARACTERIZATION
title_short Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus
title_full Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus
title_fullStr Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus
title_full_unstemmed Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus
title_sort Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus
dc.creator.none.fl_str_mv Tomás, Gonzalo
Hernández, Martín
Marandino, Ana
Panzera, Yanina
Maya, Leticia
Hernández, Diego
Pereda, Ariel Julián
Banda, Alejandro
Villegas, Pedro
Aguirre, Sebastian
Pérez, Ruben
author Tomás, Gonzalo
author_facet Tomás, Gonzalo
Hernández, Martín
Marandino, Ana
Panzera, Yanina
Maya, Leticia
Hernández, Diego
Pereda, Ariel Julián
Banda, Alejandro
Villegas, Pedro
Aguirre, Sebastian
Pérez, Ruben
author_role author
author2 Hernández, Martín
Marandino, Ana
Panzera, Yanina
Maya, Leticia
Hernández, Diego
Pereda, Ariel Julián
Banda, Alejandro
Villegas, Pedro
Aguirre, Sebastian
Pérez, Ruben
author2_role author
author
author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv DIAGNOSIS
IBDV
INFECTIOUS BURSAL DISEASE (GUMBORO)
REAL-TIME PCR
STRAIN CHARACTERIZATION
topic DIAGNOSIS
IBDV
INFECTIOUS BURSAL DISEASE (GUMBORO)
REAL-TIME PCR
STRAIN CHARACTERIZATION
purl_subject.fl_str_mv https://purl.org/becyt/ford/4.3
https://purl.org/becyt/ford/4
dc.description.none.fl_txt_mv Rapid and reliable detection and classification of infectious bursal disease viruses (IBDVs) is of crucial importance for disease surveillance and control. This study presents the development and validation of a real-time RT-PCR assay to detect and discriminate very virulent (vv) from non-vv (classic and variant) IBDV strains. The assay uses two fluorogenic, minor groove-binding (MGB) TaqMan probes targeted to a single nucleotide polymorphism (SNP) embedded in a highly conserved genomic region. The analyti- cal sensitivity of the assay was determined using serial dilutions of in vitro-transcribed RNA. The assay demonstrated a wide dynamic range between 102 and 108 standard RNA copies per reaction. Good repro- ducibility was also detected, with intra- and inter-assay coefficients of variation ranging from 0.13% to 2.23% and 0.26% to 1.92%, respectively. The assay detected successfully all the assessed vv, classical, and variant field and vaccine strains and correctly discriminated all vvIBDV strains from non-vvIBDV strains. Other common avian RNA viruses tested negative, indicating high specificity of the assay. The high sensi- tivity, rapidity, reproducibility, and specificity of the real-time RT-PCR assay make this method suitable for general and genotype-specific detection and quantitation.
Fil: Tomás, Gonzalo. Universidad del Uruguay. Facultad de Ciencias; Uruguay
Fil: Hernández, Martín. Universidad del Uruguay. Facultad de Ciencias; Uruguay
Fil: Marandino, Ana. Universidad del Uruguay. Facultad de Ciencias; Uruguay
Fil: Panzera, Yanina. Universidad de la República. Facultad de Ciencias; Uruguay
Fil: Maya, Leticia. Universidad de la República. Facultad de Ciencias; Uruguay
Fil: Hernández, Diego. Universidad de la República. Facultad de Ciencias; Uruguay
Fil: Pereda, Ariel Julián. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Banda, Alejandro. Mississippi State University; Estados Unidos
Fil: Villegas, Pedro. Georgia State University; Estados Unidos
Fil: Aguirre, Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencia y Tecnología "Dr. César Milstein". Fundación Pablo Cassará. Instituto de Ciencia y Tecnología "Dr. César Milstein"; Argentina
Fil: Pérez, Ruben. Universidad de la República. Facultad de Ciencias; Uruguay
description Rapid and reliable detection and classification of infectious bursal disease viruses (IBDVs) is of crucial importance for disease surveillance and control. This study presents the development and validation of a real-time RT-PCR assay to detect and discriminate very virulent (vv) from non-vv (classic and variant) IBDV strains. The assay uses two fluorogenic, minor groove-binding (MGB) TaqMan probes targeted to a single nucleotide polymorphism (SNP) embedded in a highly conserved genomic region. The analyti- cal sensitivity of the assay was determined using serial dilutions of in vitro-transcribed RNA. The assay demonstrated a wide dynamic range between 102 and 108 standard RNA copies per reaction. Good repro- ducibility was also detected, with intra- and inter-assay coefficients of variation ranging from 0.13% to 2.23% and 0.26% to 1.92%, respectively. The assay detected successfully all the assessed vv, classical, and variant field and vaccine strains and correctly discriminated all vvIBDV strains from non-vvIBDV strains. Other common avian RNA viruses tested negative, indicating high specificity of the assay. The high sensi- tivity, rapidity, reproducibility, and specificity of the real-time RT-PCR assay make this method suitable for general and genotype-specific detection and quantitation.
publishDate 2012
dc.date.none.fl_str_mv 2012-10
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/197963
Tomás, Gonzalo; Hernández, Martín; Marandino, Ana; Panzera, Yanina; Maya, Leticia; et al.; Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus; Elsevier Science; Journal of Virological Methods; 185; 1; 10-2012; 101-107
0166-0934
CONICET Digital
CONICET
url http://hdl.handle.net/11336/197963
identifier_str_mv Tomás, Gonzalo; Hernández, Martín; Marandino, Ana; Panzera, Yanina; Maya, Leticia; et al.; Development and validation of a TaqMan-MGB real-time RT-PCR assay for simultaneous detection and characterization of infectious bursal disease virus; Elsevier Science; Journal of Virological Methods; 185; 1; 10-2012; 101-107
0166-0934
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S0166093412002108
info:eu-repo/semantics/altIdentifier/doi/10.1016/j.jviromet.2012.06.012
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Elsevier Science
publisher.none.fl_str_mv Elsevier Science
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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