Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application

[EN] Citrus viroid V (CVd-V), a member of the species Apscaviroid epsiloncitri (family Pospiviroidae), is a graft-transmitted pathogen that spreads through infected propagation material and has been reported in several countries. Recently, it has been detected for the first time in Italy. Although C...

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Autores: Agro, Graziella, Licciardello, Grazia, Scuderi, Giuseppe, Catara, Antonino, Caruso, Andrea Giovanni, Panno, Stefano, Matic, Slavica, Davino, Salvatore, Sánchez-Navarro, Jesús-Ángel|||0000-0002-3320-2827, Pallás Benet, Vicente|||0000-0003-4954-989X
Tipo de recurso: artículo
Fecha de publicación:2026
País:España
Institución:Universitat Politècnica de València (UPV)
Repositorio:RiuNet. Repositorio Institucional de la Universitat Politécnica de Valéncia
Idioma:inglés
OAI Identifier:oai:dnet:riunet______::8d1fe758956264b885842115a8e21e8f
Acceso en línea:https://riunet.upv.es/handle/10251/234621
Access Level:acceso abierto
Palabra clave:CVd-V
RT-qPCR
Viroid
Citrus
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spelling Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field ApplicationAgro, GraziellaLicciardello, GraziaScuderi, GiuseppeCatara, AntoninoCaruso, Andrea GiovanniPanno, StefanoMatic, SlavicaDavino, SalvatoreSánchez-Navarro, Jesús-Ángel|||0000-0002-3320-2827Pallás Benet, Vicente|||0000-0003-4954-989XCVd-VRT-qPCRViroidCitrus[EN] Citrus viroid V (CVd-V), a member of the species Apscaviroid epsiloncitri (family Pospiviroidae), is a graft-transmitted pathogen that spreads through infected propagation material and has been reported in several countries. Recently, it has been detected for the first time in Italy. Although CVd-V is generally considered asymptomatic, foliar and stem symptoms have been observed in Etrog citron. In several citrus species, mixed infections of CVd-V with other viroids give synergistic effects that cause more severe symptoms. To evaluate its spread, a specific and sensitive RT-qPCR assay was developed. The assay specificity, sensitivity, and performance were compared with a conventional end-point RT-PCR, showing a higher sensitivity and no cross-reactivity with other citrus viroids. Furthermore, the assay was evaluated using two RNA extraction procedures: total RNA extraction with commercial kits and a rapid crude extract preparation using a simple extraction buffer. Moreover, RNA extracts of 111 samples, collected from commercial citrus orchards across the main Sicilian citrus-producing areas and from old varieties maintained in germplasm collection, were analyzed. The RT-qPCR revealed an overall CVd-V incidence of 8.1%. Notably, the crude extract preparation showed a sensitivity comparable with conventional total RNA extraction, with substantial savings in cost and processing time. This finding paves the way for using the developed assay with portable qPCR instruments directly in the field, as well as for routine surveillance analyses in citrus production systems.This research was funded by the SiciliAn MicronanOTecH Research and Innovation CEnter SAMOTHRACE (MUR, PNRRM4C2), spoke 3-University of Palermo S2-COMMs- Micro and Nanotechnologies for Smart and Sustainable Communities, grant number ECS_00000022.MDPI AGInstituto Universitario Mixto de Biología Molecular y Celular de PlantasMinistero dell'Istruzione, dell'Università e della RicercaRepositorio Institucional de la Universitat Politècnica de València Riunet20262026-03-01journal articlehttp://purl.org/coar/resource_type/c_6501VoRhttp://purl.org/coar/version/c_970fb48d4fbd8a85info:eu-repo/semantics/articleapplication/pdfhttps://riunet.upv.es/handle/10251/234621reponame:RiuNet. Repositorio Institucional de la Universitat Politécnica de Valénciainstname:Universitat Politècnica de València (UPV)InglésengMinistero dell'Istruzione dell'Università e della Ricerca, Italia https://doi.org/10.13039/501100003407 ECS_00000022open accesshttp://purl.org/coar/access_right/c_abf2Reconocimiento (by)http://creativecommons.org/licenses/by/4.0/info:eu-repo/semantics/openAccessoai:dnet:riunet______::8d1fe758956264b885842115a8e21e8f2026-06-13T07:49:27Z
dc.title.none.fl_str_mv Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application
title Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application
spellingShingle Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application
Agro, Graziella
CVd-V
RT-qPCR
Viroid
Citrus
title_short Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application
title_full Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application
title_fullStr Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application
title_full_unstemmed Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application
title_sort Development of a Rapid and Sensitive RT-qPCR for the Specific Detection of Citrus Viroid V and Its Field Application
dc.creator.none.fl_str_mv Agro, Graziella
Licciardello, Grazia
Scuderi, Giuseppe
Catara, Antonino
Caruso, Andrea Giovanni
Panno, Stefano
Matic, Slavica
Davino, Salvatore
Sánchez-Navarro, Jesús-Ángel|||0000-0002-3320-2827
Pallás Benet, Vicente|||0000-0003-4954-989X
author Agro, Graziella
author_facet Agro, Graziella
Licciardello, Grazia
Scuderi, Giuseppe
Catara, Antonino
Caruso, Andrea Giovanni
Panno, Stefano
Matic, Slavica
Davino, Salvatore
Sánchez-Navarro, Jesús-Ángel|||0000-0002-3320-2827
Pallás Benet, Vicente|||0000-0003-4954-989X
author_role author
author2 Licciardello, Grazia
Scuderi, Giuseppe
Catara, Antonino
Caruso, Andrea Giovanni
Panno, Stefano
Matic, Slavica
Davino, Salvatore
Sánchez-Navarro, Jesús-Ángel|||0000-0002-3320-2827
Pallás Benet, Vicente|||0000-0003-4954-989X
author2_role author
author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Instituto Universitario Mixto de Biología Molecular y Celular de Plantas
Ministero dell'Istruzione, dell'Università e della Ricerca
Repositorio Institucional de la Universitat Politècnica de València Riunet
dc.subject.none.fl_str_mv CVd-V
RT-qPCR
Viroid
Citrus
topic CVd-V
RT-qPCR
Viroid
Citrus
description [EN] Citrus viroid V (CVd-V), a member of the species Apscaviroid epsiloncitri (family Pospiviroidae), is a graft-transmitted pathogen that spreads through infected propagation material and has been reported in several countries. Recently, it has been detected for the first time in Italy. Although CVd-V is generally considered asymptomatic, foliar and stem symptoms have been observed in Etrog citron. In several citrus species, mixed infections of CVd-V with other viroids give synergistic effects that cause more severe symptoms. To evaluate its spread, a specific and sensitive RT-qPCR assay was developed. The assay specificity, sensitivity, and performance were compared with a conventional end-point RT-PCR, showing a higher sensitivity and no cross-reactivity with other citrus viroids. Furthermore, the assay was evaluated using two RNA extraction procedures: total RNA extraction with commercial kits and a rapid crude extract preparation using a simple extraction buffer. Moreover, RNA extracts of 111 samples, collected from commercial citrus orchards across the main Sicilian citrus-producing areas and from old varieties maintained in germplasm collection, were analyzed. The RT-qPCR revealed an overall CVd-V incidence of 8.1%. Notably, the crude extract preparation showed a sensitivity comparable with conventional total RNA extraction, with substantial savings in cost and processing time. This finding paves the way for using the developed assay with portable qPCR instruments directly in the field, as well as for routine surveillance analyses in citrus production systems.
publishDate 2026
dc.date.none.fl_str_mv 2026
2026-03-01
dc.type.none.fl_str_mv journal article
http://purl.org/coar/resource_type/c_6501
VoR
http://purl.org/coar/version/c_970fb48d4fbd8a85
dc.type.openaire.fl_str_mv info:eu-repo/semantics/article
format article
dc.identifier.none.fl_str_mv https://riunet.upv.es/handle/10251/234621
url https://riunet.upv.es/handle/10251/234621
dc.language.none.fl_str_mv Inglés
eng
language_invalid_str_mv Inglés
language eng
dc.relation.none.fl_str_mv Ministero dell'Istruzione dell'Università e della Ricerca, Italia https://doi.org/10.13039/501100003407 ECS_00000022
dc.rights.none.fl_str_mv open access
http://purl.org/coar/access_right/c_abf2
Reconocimiento (by)
http://creativecommons.org/licenses/by/4.0/
dc.rights.openaire.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv open access
http://purl.org/coar/access_right/c_abf2
Reconocimiento (by)
http://creativecommons.org/licenses/by/4.0/
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv MDPI AG
publisher.none.fl_str_mv MDPI AG
dc.source.none.fl_str_mv reponame:RiuNet. Repositorio Institucional de la Universitat Politécnica de Valéncia
instname:Universitat Politècnica de València (UPV)
instname_str Universitat Politècnica de València (UPV)
reponame_str RiuNet. Repositorio Institucional de la Universitat Politécnica de Valéncia
collection RiuNet. Repositorio Institucional de la Universitat Politécnica de Valéncia
repository.name.fl_str_mv
repository.mail.fl_str_mv
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score 15,81155

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