A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis
Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) are antigenically and genetically similar. The aim of this study was to develop a simple and reliable one-step real time PCR assay with high resolution melting (HRM) analysis for the simultaneous detection and differentiation of BoHV-1 and BoHV-5. O...
| Autores principales: | , , , , , , , |
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| Formato: | Artículo |
| Lenguaje: | Inglés |
| Publicado: |
Elsevier
2019
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| Materias: | |
| Acceso en línea: | https://www.sciencedirect.com/science/article/pii/S0166093415003390 http://hdl.handle.net/20.500.12123/4981 https://doi.org/10.1016/j.jviromet.2015.10.005 |
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| author | Marin, Maia Solange Quintana, Silvina Leunda, Maria Rosa Recavarren, Mariana Ines Pagnuco, Inti Anabela Spath, Ernesto Juan Perez, Sandra Odeon, Anselmo Carlos |
| author_browse | Leunda, Maria Rosa Marin, Maia Solange Odeon, Anselmo Carlos Pagnuco, Inti Anabela Perez, Sandra Quintana, Silvina Recavarren, Mariana Ines Spath, Ernesto Juan |
| author_facet | Marin, Maia Solange Quintana, Silvina Leunda, Maria Rosa Recavarren, Mariana Ines Pagnuco, Inti Anabela Spath, Ernesto Juan Perez, Sandra Odeon, Anselmo Carlos |
| author_sort | Marin, Maia Solange |
| collection | INTA Digital |
| description | Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) are antigenically and genetically similar. The aim of this study was to develop a simple and reliable one-step real time PCR assay with high resolution melting (HRM) analysis for the simultaneous detection and differentiation of BoHV-1 and BoHV-5. Optimization of assay conditions was performed with DNA from reference strains. Then, DNA from field isolates, clinical samples and tissue samples of experimentally infected animals were studied by real time PCR-HRM. An efficient amplification of real time PCR products was obtained, and a clear melting curve and appropriate melting peaks for both viruses were achieved in the HRM curve analysis for BoHV type identification. BoHV was identified in all of the isolates and clinical samples, and BoHV types were properly differentiated. Furthermore, viral DNA was detected in 12/18 and 7/18 samples from BoHV-1- and BoHV-5-infected calves, respectively. Real time PCR-HRM achieved a higher sensitivity compared with virus isolation or conventional PCR. In this study, HRM was used as a novel procedure. This method provides rapid, sensitive, specific and simultaneous detection of bovine alpha-herpesviruses DNA. Thus, this technique is an excellent tool for diagnosis, research and epidemiological studies of these viruses in cattle. |
| format | Artículo |
| id | INTA4981 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2019 |
| publishDateRange | 2019 |
| publishDateSort | 2019 |
| publisher | Elsevier |
| publisherStr | Elsevier |
| record_format | dspace |
| spelling | INTA49812019-04-25T13:10:58Z A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis Marin, Maia Solange Quintana, Silvina Leunda, Maria Rosa Recavarren, Mariana Ines Pagnuco, Inti Anabela Spath, Ernesto Juan Perez, Sandra Odeon, Anselmo Carlos Ganado Bovino Enfermedades de los Animales Herpes Virus Bovino PCR Diagnóstico Cattle Animal Diseases Bovine Herpesvirus Diagnosis Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) are antigenically and genetically similar. The aim of this study was to develop a simple and reliable one-step real time PCR assay with high resolution melting (HRM) analysis for the simultaneous detection and differentiation of BoHV-1 and BoHV-5. Optimization of assay conditions was performed with DNA from reference strains. Then, DNA from field isolates, clinical samples and tissue samples of experimentally infected animals were studied by real time PCR-HRM. An efficient amplification of real time PCR products was obtained, and a clear melting curve and appropriate melting peaks for both viruses were achieved in the HRM curve analysis for BoHV type identification. BoHV was identified in all of the isolates and clinical samples, and BoHV types were properly differentiated. Furthermore, viral DNA was detected in 12/18 and 7/18 samples from BoHV-1- and BoHV-5-infected calves, respectively. Real time PCR-HRM achieved a higher sensitivity compared with virus isolation or conventional PCR. In this study, HRM was used as a novel procedure. This method provides rapid, sensitive, specific and simultaneous detection of bovine alpha-herpesviruses DNA. Thus, this technique is an excellent tool for diagnosis, research and epidemiological studies of these viruses in cattle. EEA Balcarce Fil: Marin, Maia Solange. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Quintana, Silvina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Análisis Fares Taie; Argentina Fil: Leunda, Maria Rosa. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina Fil: Recavarren, Mariana Ines. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto de Análisis Fares Taie; Argentina Fil: Pagnuco, Inti Anabela. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Spath, Ernesto Juan. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina Fil: Perez, Sandra. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; Argentina Fil: Odeón, Anselmo Carlos. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina 2019-04-25T13:08:56Z 2019-04-25T13:08:56Z 2016-01 info:ar-repo/semantics/artículo info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion https://www.sciencedirect.com/science/article/pii/S0166093415003390 http://hdl.handle.net/20.500.12123/4981 0166-0934 https://doi.org/10.1016/j.jviromet.2015.10.005 eng info:eu-repo/semantics/restrictedAccess application/pdf Elsevier Journal of Virological Methods 227 : 14-22 (January 2016) |
| spellingShingle | Ganado Bovino Enfermedades de los Animales Herpes Virus Bovino PCR Diagnóstico Cattle Animal Diseases Bovine Herpesvirus Diagnosis Marin, Maia Solange Quintana, Silvina Leunda, Maria Rosa Recavarren, Mariana Ines Pagnuco, Inti Anabela Spath, Ernesto Juan Perez, Sandra Odeon, Anselmo Carlos A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis |
| title | A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis |
| title_full | A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis |
| title_fullStr | A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis |
| title_full_unstemmed | A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis |
| title_short | A new method for simultaneous detection and discrimination of Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) using real time PCR with high resolution melting (HRM) analysis |
| title_sort | new method for simultaneous detection and discrimination of bovine herpesvirus types 1 bohv 1 and 5 bohv 5 using real time pcr with high resolution melting hrm analysis |
| topic | Ganado Bovino Enfermedades de los Animales Herpes Virus Bovino PCR Diagnóstico Cattle Animal Diseases Bovine Herpesvirus Diagnosis |
| url | https://www.sciencedirect.com/science/article/pii/S0166093415003390 http://hdl.handle.net/20.500.12123/4981 https://doi.org/10.1016/j.jviromet.2015.10.005 |
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