Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit

Rapid and reliable detection of Monilinia latent infections is needed to prevent and control dispersion of Monilinia spp. in infected localities and non-infected countries. A fast multiplex quantitative real-time PCR method (qPCR) for the detection and identification of Monilinia spp. latent infecti...

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Autores: García-Benitez, Carlos, Melgarejo, Paloma, Beniusis, A., Guinet, Cécile, Özben, S., Değirmenci, K., Valente, M. T., Riccioni, L., De Cal Cortina, Antonieta
Formato: artículo
Fecha de publicación:2017
País:España
Recursos:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/293577
Acesso em linha:http://hdl.handle.net/10261/293577
Access Level:acceso abierto
Palavra-chave:Brown rot
qPCR
Inter-laboratory validation
Performance assessment
Sensitivity
Specificity
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spelling Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruitGarcía-Benitez, CarlosMelgarejo, PalomaBeniusis, A.Guinet, CécileÖzben, S.Değirmenci, K.Valente, M. T.Riccioni, L.De Cal Cortina, AntonietaBrown rotqPCRInter-laboratory validationPerformance assessmentSensitivitySpecificityRapid and reliable detection of Monilinia latent infections is needed to prevent and control dispersion of Monilinia spp. in infected localities and non-infected countries. A fast multiplex quantitative real-time PCR method (qPCR) for the detection and identification of Monilinia spp. latent infections in blossoms and fruit of nectarine trees (Prunus persica var. nucipersica) was tested in an inter-laboratory trial. The test performance study involving five laboratories was conducted to validate the sensitivity and specificity of several real-time PCR platforms for the detection of low amounts of Monilinia DNA (latent infections), using a common protocol, and to identify possible difficulties when these tests were implemented by diagnostic laboratories or national reference centres. The method has two hydrolysis probes distinguishing between Monilinia fructicola and M. fructigena/M. laxa. Validation included test performance accuracy, analytical specificity and sensitivity, repeatability, and reproducibility, as defined by standard PM7/98 of the European Plant Protection Organization (EPPO). All qPCR platforms detected Monilinia latent infections and mycelium samples with both hydrolysis probes, and healthy flowers and fruit samples gave negative results. The method specificity was consistent between different laboratories, despite different equipment used, and there were no laboratories with z-scores in the unacceptable region. Monilinia fructicola latent infection samples were correctly detected by all laboratories, but some M. laxa samples were cross-detected as if they were M. fructicola. Monilinia laxa cross-detection could be compensated by including the allelic discrimination step in qPCR runs, which permitted differentiating between M. fructicola and M. laxa samples. The inter-laboratory comparison demonstrated the robustness of the developed method and confirmed in-house validation data. This method could be used to detect latent infections of Monilinia in asymptomatic nectarine fruit and flowers. © 2017 Author(s).Peer reviewedFirenze University PressGarcía-Benitez, Carlos [0000-0002-8456-0421]Melgarejo, Paloma [0000-0002-3698-8896]De Cal Cortina, Antonieta [0000-0002-7725-7782]Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]202320232017info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501http://hdl.handle.net/10261/293577reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)InglésDepartamento de Protección VegetalSíinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/2935772026-05-22T06:33:51Z
dc.title.none.fl_str_mv Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit
title Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit
spellingShingle Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit
García-Benitez, Carlos
Brown rot
qPCR
Inter-laboratory validation
Performance assessment
Sensitivity
Specificity
title_short Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit
title_full Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit
title_fullStr Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit
title_full_unstemmed Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit
title_sort Proficiency of real-time PCR detection of latent Monilinia spp. infection in nectarine flowers and fruit
dc.creator.none.fl_str_mv García-Benitez, Carlos
Melgarejo, Paloma
Beniusis, A.
Guinet, Cécile
Özben, S.
Değirmenci, K.
Valente, M. T.
Riccioni, L.
De Cal Cortina, Antonieta
author García-Benitez, Carlos
author_facet García-Benitez, Carlos
Melgarejo, Paloma
Beniusis, A.
Guinet, Cécile
Özben, S.
Değirmenci, K.
Valente, M. T.
Riccioni, L.
De Cal Cortina, Antonieta
author_role author
author2 Melgarejo, Paloma
Beniusis, A.
Guinet, Cécile
Özben, S.
Değirmenci, K.
Valente, M. T.
Riccioni, L.
De Cal Cortina, Antonieta
author2_role author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv García-Benitez, Carlos [0000-0002-8456-0421]
Melgarejo, Paloma [0000-0002-3698-8896]
De Cal Cortina, Antonieta [0000-0002-7725-7782]
Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv Brown rot
qPCR
Inter-laboratory validation
Performance assessment
Sensitivity
Specificity
topic Brown rot
qPCR
Inter-laboratory validation
Performance assessment
Sensitivity
Specificity
description Rapid and reliable detection of Monilinia latent infections is needed to prevent and control dispersion of Monilinia spp. in infected localities and non-infected countries. A fast multiplex quantitative real-time PCR method (qPCR) for the detection and identification of Monilinia spp. latent infections in blossoms and fruit of nectarine trees (Prunus persica var. nucipersica) was tested in an inter-laboratory trial. The test performance study involving five laboratories was conducted to validate the sensitivity and specificity of several real-time PCR platforms for the detection of low amounts of Monilinia DNA (latent infections), using a common protocol, and to identify possible difficulties when these tests were implemented by diagnostic laboratories or national reference centres. The method has two hydrolysis probes distinguishing between Monilinia fructicola and M. fructigena/M. laxa. Validation included test performance accuracy, analytical specificity and sensitivity, repeatability, and reproducibility, as defined by standard PM7/98 of the European Plant Protection Organization (EPPO). All qPCR platforms detected Monilinia latent infections and mycelium samples with both hydrolysis probes, and healthy flowers and fruit samples gave negative results. The method specificity was consistent between different laboratories, despite different equipment used, and there were no laboratories with z-scores in the unacceptable region. Monilinia fructicola latent infection samples were correctly detected by all laboratories, but some M. laxa samples were cross-detected as if they were M. fructicola. Monilinia laxa cross-detection could be compensated by including the allelic discrimination step in qPCR runs, which permitted differentiating between M. fructicola and M. laxa samples. The inter-laboratory comparison demonstrated the robustness of the developed method and confirmed in-house validation data. This method could be used to detect latent infections of Monilinia in asymptomatic nectarine fruit and flowers. © 2017 Author(s).
publishDate 2017
dc.date.none.fl_str_mv 2017
2023
2023
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
format article
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/293577
url http://hdl.handle.net/10261/293577
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv Departamento de Protección Vegetal
Sí
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv Firenze University Press
publisher.none.fl_str_mv Firenze University Press
dc.source.none.fl_str_mv reponame:DIGITAL.CSIC. Repositorio Institucional del CSIC
instname:Consejo Superior de Investigaciones Científicas (CSIC)
instname_str Consejo Superior de Investigaciones Científicas (CSIC)
reponame_str DIGITAL.CSIC. Repositorio Institucional del CSIC
collection DIGITAL.CSIC. Repositorio Institucional del CSIC
repository.name.fl_str_mv
repository.mail.fl_str_mv
_version_ 1869416564591165440
score 15,812429

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