Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA

Autores
de Francesco, Agustina; Acosta, Norma Rebeca; Maria Inés Plata; Garcia, Maria Laura
Año de publicación
2015
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Citrus is one of the most economically important fruit crops in the world. Citrus psorosis is a serious disease affecting mainly oranges and mandarins in Argentina and Uruguay. The causal agent is Citrus psorosis virus (CPsV), an ophiovirus with a tripartite ssRNA genome of negative polarity. The coat protein (CP), the most abundant viral protein in infected plants, has been used to detect CPsV by TAS‐ELISA, but only biological indexing, requiring 1 year, is the current and validated technique for diagnosis of citrus psorosis. In this study, a SYBR Green RT‐qPCR protocol was developed, with primers designed to the most conserved region of the cp gene. We tested their specificity and sensitivity in comparison with TAS‐ELISA. This RT‐qPCR was applied successfully to field samples from Argentina, to a variety of isolates from different countries maintained in the greenhouse, to young seedlings and old trees from a psorosis natural transmission plot, and to transgenic citrus expressing the cp gene of CPsV or a fragment thereof. This method allowed accurate quantification of viral titer and cp gene expression in transgenic plants, which could not be detected previously. The sensitivity and reliability of quantitative CPsV detection were improved with greater speed using commercial reagents, and the sensitivity was three orders of magnitude higher than that of TAS‐ELISA. All these data encourage its validation.
Fil: de Francesco, Agustina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina
Fil: Acosta, Norma Rebeca. Estación Experimental Agropecuaria Concordia; Argentina
Fil: Maria Inés Plata. Estación Experimental Agropecuaria Concordia; Argentina
Fil: Garcia, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina
Materia
Citrus Psorosis Virus
Diagnosis
Rt-Qpcr
Tas-Elisa
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:11336/47890
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/47890
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISAde Francesco, AgustinaAcosta, Norma RebecaMaria Inés PlataGarcia, Maria LauraCitrus Psorosis VirusDiagnosisRt-QpcrTas-Elisahttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Citrus is one of the most economically important fruit crops in the world. Citrus psorosis is a serious disease affecting mainly oranges and mandarins in Argentina and Uruguay. The causal agent is Citrus psorosis virus (CPsV), an ophiovirus with a tripartite ssRNA genome of negative polarity. The coat protein (CP), the most abundant viral protein in infected plants, has been used to detect CPsV by TAS‐ELISA, but only biological indexing, requiring 1 year, is the current and validated technique for diagnosis of citrus psorosis. In this study, a SYBR Green RT‐qPCR protocol was developed, with primers designed to the most conserved region of the cp gene. We tested their specificity and sensitivity in comparison with TAS‐ELISA. This RT‐qPCR was applied successfully to field samples from Argentina, to a variety of isolates from different countries maintained in the greenhouse, to young seedlings and old trees from a psorosis natural transmission plot, and to transgenic citrus expressing the cp gene of CPsV or a fragment thereof. This method allowed accurate quantification of viral titer and cp gene expression in transgenic plants, which could not be detected previously. The sensitivity and reliability of quantitative CPsV detection were improved with greater speed using commercial reagents, and the sensitivity was three orders of magnitude higher than that of TAS‐ELISA. All these data encourage its validation.Fil: de Francesco, Agustina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Acosta, Norma Rebeca. Estación Experimental Agropecuaria Concordia; ArgentinaFil: Maria Inés Plata. Estación Experimental Agropecuaria Concordia; ArgentinaFil: Garcia, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaWiley Blackwell Publishing, Inc2015-12info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/47890de Francesco, Agustina; Acosta, Norma Rebeca; Maria Inés Plata; Garcia, Maria Laura; Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA; Wiley Blackwell Publishing, Inc; Journal of Phytopathology-Phytopathologische Zeitschrift; 163; 11-12; 12-2015; 915-9250931-1785CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1111/jph.12392info:eu-repo/semantics/altIdentifier/url/https://onlinelibrary.wiley.com/doi/abs/10.1111/jph.12392info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-10T13:17:47Zoai:ri.conicet.gov.ar:11336/47890instacron: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:34982025-09-10 13:17:47.588CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA
title Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA
spellingShingle Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA
de Francesco, Agustina
Citrus Psorosis Virus
Diagnosis
Rt-Qpcr
Tas-Elisa
title_short Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA
title_full Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA
title_fullStr Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA
title_full_unstemmed Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA
title_sort Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA
dc.creator.none.fl_str_mv de Francesco, Agustina
Acosta, Norma Rebeca
Maria Inés Plata
Garcia, Maria Laura
author de Francesco, Agustina
author_facet de Francesco, Agustina
Acosta, Norma Rebeca
Maria Inés Plata
Garcia, Maria Laura
author_role author
author2 Acosta, Norma Rebeca
Maria Inés Plata
Garcia, Maria Laura
author2_role author
author
author
dc.subject.none.fl_str_mv Citrus Psorosis Virus
Diagnosis
Rt-Qpcr
Tas-Elisa
topic Citrus Psorosis Virus
Diagnosis
Rt-Qpcr
Tas-Elisa
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Citrus is one of the most economically important fruit crops in the world. Citrus psorosis is a serious disease affecting mainly oranges and mandarins in Argentina and Uruguay. The causal agent is Citrus psorosis virus (CPsV), an ophiovirus with a tripartite ssRNA genome of negative polarity. The coat protein (CP), the most abundant viral protein in infected plants, has been used to detect CPsV by TAS‐ELISA, but only biological indexing, requiring 1 year, is the current and validated technique for diagnosis of citrus psorosis. In this study, a SYBR Green RT‐qPCR protocol was developed, with primers designed to the most conserved region of the cp gene. We tested their specificity and sensitivity in comparison with TAS‐ELISA. This RT‐qPCR was applied successfully to field samples from Argentina, to a variety of isolates from different countries maintained in the greenhouse, to young seedlings and old trees from a psorosis natural transmission plot, and to transgenic citrus expressing the cp gene of CPsV or a fragment thereof. This method allowed accurate quantification of viral titer and cp gene expression in transgenic plants, which could not be detected previously. The sensitivity and reliability of quantitative CPsV detection were improved with greater speed using commercial reagents, and the sensitivity was three orders of magnitude higher than that of TAS‐ELISA. All these data encourage its validation.
Fil: de Francesco, Agustina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina
Fil: Acosta, Norma Rebeca. Estación Experimental Agropecuaria Concordia; Argentina
Fil: Maria Inés Plata. Estación Experimental Agropecuaria Concordia; Argentina
Fil: Garcia, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentina
description Citrus is one of the most economically important fruit crops in the world. Citrus psorosis is a serious disease affecting mainly oranges and mandarins in Argentina and Uruguay. The causal agent is Citrus psorosis virus (CPsV), an ophiovirus with a tripartite ssRNA genome of negative polarity. The coat protein (CP), the most abundant viral protein in infected plants, has been used to detect CPsV by TAS‐ELISA, but only biological indexing, requiring 1 year, is the current and validated technique for diagnosis of citrus psorosis. In this study, a SYBR Green RT‐qPCR protocol was developed, with primers designed to the most conserved region of the cp gene. We tested their specificity and sensitivity in comparison with TAS‐ELISA. This RT‐qPCR was applied successfully to field samples from Argentina, to a variety of isolates from different countries maintained in the greenhouse, to young seedlings and old trees from a psorosis natural transmission plot, and to transgenic citrus expressing the cp gene of CPsV or a fragment thereof. This method allowed accurate quantification of viral titer and cp gene expression in transgenic plants, which could not be detected previously. The sensitivity and reliability of quantitative CPsV detection were improved with greater speed using commercial reagents, and the sensitivity was three orders of magnitude higher than that of TAS‐ELISA. All these data encourage its validation.
publishDate 2015
dc.date.none.fl_str_mv 2015-12
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/11336/47890
de Francesco, Agustina; Acosta, Norma Rebeca; Maria Inés Plata; Garcia, Maria Laura; Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA; Wiley Blackwell Publishing, Inc; Journal of Phytopathology-Phytopathologische Zeitschrift; 163; 11-12; 12-2015; 915-925
0931-1785
CONICET Digital
CONICET
url http://hdl.handle.net/11336/47890
identifier_str_mv de Francesco, Agustina; Acosta, Norma Rebeca; Maria Inés Plata; Garcia, Maria Laura; Improved Detection of Citrus psorosis virus and Coat Protein-Derived Transgenes in Citrus Plants: Comparison Between RT-qPCR and TAS-ELISA; Wiley Blackwell Publishing, Inc; Journal of Phytopathology-Phytopathologische Zeitschrift; 163; 11-12; 12-2015; 915-925
0931-1785
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1111/jph.12392
info:eu-repo/semantics/altIdentifier/url/https://onlinelibrary.wiley.com/doi/abs/10.1111/jph.12392
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
dc.publisher.none.fl_str_mv Wiley Blackwell Publishing, Inc
publisher.none.fl_str_mv Wiley Blackwell Publishing, Inc
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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score 12.993085

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