Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates

Autores
Maidana, Silvina Soledad; Morano, Cintia Débora; Cianfrini, Daniela; Campos, Fabrício Souza; Roehe, Paulo Michel; Siedler, Bianca; De Stefano, Gabriel Alejandro; Mauroy, Axel; Thiry, Etienne; Romera, Sonia Alejandra
Año de publicación
2013
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Background: Several types and subtypes of bovine herpesviruses 1 and 5 (BoHV-1 and BoHV-5) have been associated to different clinical conditions of cattle, making type/subtype differentiation essential to understand the pathogenesis and epidemiology of BoHV infections. BoHV-5 subtyping is currently carried out by BstEII restriction enzyme analysis (REA) of the complete virus genome. This method allowed the description of three subtypes, one of which is the most widespread while the remaining two have so far only been found in South America. The present work describes a multiplex PCR followed by REA for BoHV-5 subtyping. Results: The method consists in the simultaneous amplification of glycoprotein B and UL54 gene fragments of 534 and 669 base pairs (bp), respectively, BstEII digestion of amplicons, separation of products in 1% agarose gels, and analysis of fragment length polymorphims. The multiplex PCR detected up to 227 BoHV-5 genome copies and 9.2 × 105 BoHV-5 genome copies when DNA was extracted from purified virus or infected tissue homogenates, respectively. The applicability of multiplex PCR-REA was demonstrated on 3 BoHV-5 reference strains. In addition, subtyping of two new isolates and seventeen previously reported ones (17 BHV-5a and 2 BHV-5b) by this method gave coincident results with those obtained with the classic BstEII REA assay. Conclusions: Multiplex PCR-REA provides a new tool for the fast and simple diagnosis and subtyping of BoHV-5.
Instituto de Virología
Fil: Maidana, Silvina Soledad. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Morano, Cintia Débora Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina.
Fil: Cianfrini, Daniela. Tecnovax SA; Argentina
Fil: Campos, Fabrício Souza. Universidade Federal do Rio Grande do Sul. Institute of Basic Health Sciences. Department of Microbiology, Immunology and Parasitology. Virology Laboratory; Brasil
Fil: Roehe, Paulo Michel. Universidade Federal do Rio Grande do Sul. Institute of Basic Health Sciences. Department of Microbiology, Immunology and Parasitology. Virology Laboratory; Brasil
Fil: Siedler, Bianca. Universidade Federal de Pelotas. Laboratório de Bioprocessos; Brasil
Fil: De Stefano, Gabriel Alejandro. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina
Fil: Mauroy, Axel. University of Liège. Fundamental and Applied Research on Animal Health Center and Faculty of Veterinary Medicine. Veterinary Virology and Animal Viral Diseases; Bélgica
Fil: Thiry, Etienne. University of Liège. Fundamental and Applied Research on Animal Health Center and Faculty of Veterinary Medicine. Veterinary Virology and Animal Viral Diseases; Bélgica
Fil: Romera, Sonia Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad del Salvador. Cátedra de Inmunología; Argentina
Fuente
BMC Veterinary Research 39 : 111 (2013)
Materia
Enfermedades de los Animales
Ganado Bovino
Virus de los Animales
Herpesviridae
PCR
Polimorfismo
Herpes Virus Bovino
Animal Diseases
Cattle
Animal Viruses
Polymorphism
Bovine Herpesvirus
Herpesvirus
Reacción en Cadena de la Polimerasa
Nivel de accesibilidad
acceso abierto
Condiciones de uso
http://creativecommons.org/licenses/by-nc-sa/4.0/
Repositorio
INTA Digital (INTA)
Institución
Instituto Nacional de Tecnología Agropecuaria
OAI Identificador
oai:localhost:20.500.12123/4276
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oai_identifier_str oai:localhost:20.500.12123/4276
network_acronym_str INTADig
repository_id_str l
network_name_str INTA Digital (INTA)
spelling Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolatesMaidana, Silvina SoledadMorano, Cintia DéboraCianfrini, DanielaCampos, Fabrício SouzaRoehe, Paulo MichelSiedler, BiancaDe Stefano, Gabriel AlejandroMauroy, AxelThiry, EtienneRomera, Sonia AlejandraEnfermedades de los AnimalesGanado BovinoVirus de los AnimalesHerpesviridaePCRPolimorfismoHerpes Virus BovinoAnimal DiseasesCattleAnimal VirusesPolymorphismBovine HerpesvirusHerpesvirusReacción en Cadena de la PolimerasaBackground: Several types and subtypes of bovine herpesviruses 1 and 5 (BoHV-1 and BoHV-5) have been associated to different clinical conditions of cattle, making type/subtype differentiation essential to understand the pathogenesis and epidemiology of BoHV infections. BoHV-5 subtyping is currently carried out by BstEII restriction enzyme analysis (REA) of the complete virus genome. This method allowed the description of three subtypes, one of which is the most widespread while the remaining two have so far only been found in South America. The present work describes a multiplex PCR followed by REA for BoHV-5 subtyping. Results: The method consists in the simultaneous amplification of glycoprotein B and UL54 gene fragments of 534 and 669 base pairs (bp), respectively, BstEII digestion of amplicons, separation of products in 1% agarose gels, and analysis of fragment length polymorphims. The multiplex PCR detected up to 227 BoHV-5 genome copies and 9.2 × 105 BoHV-5 genome copies when DNA was extracted from purified virus or infected tissue homogenates, respectively. The applicability of multiplex PCR-REA was demonstrated on 3 BoHV-5 reference strains. In addition, subtyping of two new isolates and seventeen previously reported ones (17 BHV-5a and 2 BHV-5b) by this method gave coincident results with those obtained with the classic BstEII REA assay. Conclusions: Multiplex PCR-REA provides a new tool for the fast and simple diagnosis and subtyping of BoHV-5.Instituto de VirologíaFil: Maidana, Silvina Soledad. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Morano, Cintia Débora Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina.Fil: Cianfrini, Daniela. Tecnovax SA; ArgentinaFil: Campos, Fabrício Souza. Universidade Federal do Rio Grande do Sul. Institute of Basic Health Sciences. Department of Microbiology, Immunology and Parasitology. Virology Laboratory; BrasilFil: Roehe, Paulo Michel. Universidade Federal do Rio Grande do Sul. Institute of Basic Health Sciences. Department of Microbiology, Immunology and Parasitology. Virology Laboratory; BrasilFil: Siedler, Bianca. Universidade Federal de Pelotas. Laboratório de Bioprocessos; BrasilFil: De Stefano, Gabriel Alejandro. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; ArgentinaFil: Mauroy, Axel. University of Liège. Fundamental and Applied Research on Animal Health Center and Faculty of Veterinary Medicine. Veterinary Virology and Animal Viral Diseases; BélgicaFil: Thiry, Etienne. University of Liège. Fundamental and Applied Research on Animal Health Center and Faculty of Veterinary Medicine. Veterinary Virology and Animal Viral Diseases; BélgicaFil: Romera, Sonia Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad del Salvador. Cátedra de Inmunología; ArgentinaBMC2019-01-16T15:24:20Z2019-01-16T15:24:20Z2013-06info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfhttps://bmcvetres.biomedcentral.com/articles/10.1186/1746-6148-9-111http://hdl.handle.net/20.500.12123/42761746-6148https://doi.org/10.1186/1746-6148-9-111BMC Veterinary Research 39 : 111 (2013)reponame:INTA Digital (INTA)instname:Instituto Nacional de Tecnología Agropecuariaenginfo:eu-repo/semantics/openAccesshttp://creativecommons.org/licenses/by-nc-sa/4.0/Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)2025-09-29T13:44:33Zoai:localhost:20.500.12123/4276instacron:INTAInstitucionalhttp://repositorio.inta.gob.ar/Organismo científico-tecnológicoNo correspondehttp://repositorio.inta.gob.ar/oai/requesttripaldi.nicolas@inta.gob.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:l2025-09-29 13:44:33.455INTA Digital (INTA) - Instituto Nacional de Tecnología Agropecuariafalse
dc.title.none.fl_str_mv Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates
title Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates
spellingShingle Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates
Maidana, Silvina Soledad
Enfermedades de los Animales
Ganado Bovino
Virus de los Animales
Herpesviridae
PCR
Polimorfismo
Herpes Virus Bovino
Animal Diseases
Cattle
Animal Viruses
Polymorphism
Bovine Herpesvirus
Herpesvirus
Reacción en Cadena de la Polimerasa
title_short Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates
title_full Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates
title_fullStr Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates
title_full_unstemmed Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates
title_sort Multiplex PCR followed by restriction length polymorphism analysis for the subtyping of bovine herpesvirus 5 isolates
dc.creator.none.fl_str_mv Maidana, Silvina Soledad
Morano, Cintia Débora
Cianfrini, Daniela
Campos, Fabrício Souza
Roehe, Paulo Michel
Siedler, Bianca
De Stefano, Gabriel Alejandro
Mauroy, Axel
Thiry, Etienne
Romera, Sonia Alejandra
author Maidana, Silvina Soledad
author_facet Maidana, Silvina Soledad
Morano, Cintia Débora
Cianfrini, Daniela
Campos, Fabrício Souza
Roehe, Paulo Michel
Siedler, Bianca
De Stefano, Gabriel Alejandro
Mauroy, Axel
Thiry, Etienne
Romera, Sonia Alejandra
author_role author
author2 Morano, Cintia Débora
Cianfrini, Daniela
Campos, Fabrício Souza
Roehe, Paulo Michel
Siedler, Bianca
De Stefano, Gabriel Alejandro
Mauroy, Axel
Thiry, Etienne
Romera, Sonia Alejandra
author2_role author
author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Enfermedades de los Animales
Ganado Bovino
Virus de los Animales
Herpesviridae
PCR
Polimorfismo
Herpes Virus Bovino
Animal Diseases
Cattle
Animal Viruses
Polymorphism
Bovine Herpesvirus
Herpesvirus
Reacción en Cadena de la Polimerasa
topic Enfermedades de los Animales
Ganado Bovino
Virus de los Animales
Herpesviridae
PCR
Polimorfismo
Herpes Virus Bovino
Animal Diseases
Cattle
Animal Viruses
Polymorphism
Bovine Herpesvirus
Herpesvirus
Reacción en Cadena de la Polimerasa
dc.description.none.fl_txt_mv Background: Several types and subtypes of bovine herpesviruses 1 and 5 (BoHV-1 and BoHV-5) have been associated to different clinical conditions of cattle, making type/subtype differentiation essential to understand the pathogenesis and epidemiology of BoHV infections. BoHV-5 subtyping is currently carried out by BstEII restriction enzyme analysis (REA) of the complete virus genome. This method allowed the description of three subtypes, one of which is the most widespread while the remaining two have so far only been found in South America. The present work describes a multiplex PCR followed by REA for BoHV-5 subtyping. Results: The method consists in the simultaneous amplification of glycoprotein B and UL54 gene fragments of 534 and 669 base pairs (bp), respectively, BstEII digestion of amplicons, separation of products in 1% agarose gels, and analysis of fragment length polymorphims. The multiplex PCR detected up to 227 BoHV-5 genome copies and 9.2 × 105 BoHV-5 genome copies when DNA was extracted from purified virus or infected tissue homogenates, respectively. The applicability of multiplex PCR-REA was demonstrated on 3 BoHV-5 reference strains. In addition, subtyping of two new isolates and seventeen previously reported ones (17 BHV-5a and 2 BHV-5b) by this method gave coincident results with those obtained with the classic BstEII REA assay. Conclusions: Multiplex PCR-REA provides a new tool for the fast and simple diagnosis and subtyping of BoHV-5.
Instituto de Virología
Fil: Maidana, Silvina Soledad. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Morano, Cintia Débora Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina.
Fil: Cianfrini, Daniela. Tecnovax SA; Argentina
Fil: Campos, Fabrício Souza. Universidade Federal do Rio Grande do Sul. Institute of Basic Health Sciences. Department of Microbiology, Immunology and Parasitology. Virology Laboratory; Brasil
Fil: Roehe, Paulo Michel. Universidade Federal do Rio Grande do Sul. Institute of Basic Health Sciences. Department of Microbiology, Immunology and Parasitology. Virology Laboratory; Brasil
Fil: Siedler, Bianca. Universidade Federal de Pelotas. Laboratório de Bioprocessos; Brasil
Fil: De Stefano, Gabriel Alejandro. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina
Fil: Mauroy, Axel. University of Liège. Fundamental and Applied Research on Animal Health Center and Faculty of Veterinary Medicine. Veterinary Virology and Animal Viral Diseases; Bélgica
Fil: Thiry, Etienne. University of Liège. Fundamental and Applied Research on Animal Health Center and Faculty of Veterinary Medicine. Veterinary Virology and Animal Viral Diseases; Bélgica
Fil: Romera, Sonia Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad del Salvador. Cátedra de Inmunología; Argentina
description Background: Several types and subtypes of bovine herpesviruses 1 and 5 (BoHV-1 and BoHV-5) have been associated to different clinical conditions of cattle, making type/subtype differentiation essential to understand the pathogenesis and epidemiology of BoHV infections. BoHV-5 subtyping is currently carried out by BstEII restriction enzyme analysis (REA) of the complete virus genome. This method allowed the description of three subtypes, one of which is the most widespread while the remaining two have so far only been found in South America. The present work describes a multiplex PCR followed by REA for BoHV-5 subtyping. Results: The method consists in the simultaneous amplification of glycoprotein B and UL54 gene fragments of 534 and 669 base pairs (bp), respectively, BstEII digestion of amplicons, separation of products in 1% agarose gels, and analysis of fragment length polymorphims. The multiplex PCR detected up to 227 BoHV-5 genome copies and 9.2 × 105 BoHV-5 genome copies when DNA was extracted from purified virus or infected tissue homogenates, respectively. The applicability of multiplex PCR-REA was demonstrated on 3 BoHV-5 reference strains. In addition, subtyping of two new isolates and seventeen previously reported ones (17 BHV-5a and 2 BHV-5b) by this method gave coincident results with those obtained with the classic BstEII REA assay. Conclusions: Multiplex PCR-REA provides a new tool for the fast and simple diagnosis and subtyping of BoHV-5.
publishDate 2013
dc.date.none.fl_str_mv 2013-06
2019-01-16T15:24:20Z
2019-01-16T15:24:20Z
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 https://bmcvetres.biomedcentral.com/articles/10.1186/1746-6148-9-111
http://hdl.handle.net/20.500.12123/4276
1746-6148
https://doi.org/10.1186/1746-6148-9-111
url https://bmcvetres.biomedcentral.com/articles/10.1186/1746-6148-9-111
http://hdl.handle.net/20.500.12123/4276
https://doi.org/10.1186/1746-6148-9-111
identifier_str_mv 1746-6148
dc.language.none.fl_str_mv eng
language eng
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
http://creativecommons.org/licenses/by-nc-sa/4.0/
Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)
eu_rights_str_mv openAccess
rights_invalid_str_mv http://creativecommons.org/licenses/by-nc-sa/4.0/
Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv BMC
publisher.none.fl_str_mv BMC
dc.source.none.fl_str_mv BMC Veterinary Research 39 : 111 (2013)
reponame:INTA Digital (INTA)
instname:Instituto Nacional de Tecnología Agropecuaria
reponame_str INTA Digital (INTA)
collection INTA Digital (INTA)
instname_str Instituto Nacional de Tecnología Agropecuaria
repository.name.fl_str_mv INTA Digital (INTA) - Instituto Nacional de Tecnología Agropecuaria
repository.mail.fl_str_mv tripaldi.nicolas@inta.gob.ar
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