A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences

Autores
Iraola, Gregorio; Pérez, Ruben; Betancor, Laura; Marandino, Ana; Morsella, Claudia Graciela; Mendez, Maria Alejandra; Paolicchi, Fernando; Piccirillo, Alessandra; Tomás, Gonzalo; Velilla, Alejandra Vanesa; Calleros, Lucía
Año de publicación
2016
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Background: Campylobacter fetus is a pathogen of major concern for animal and human health. The species shows a great intraspecific variation, with three subspecies: C. fetus subsp. fetus, C. fetus subsp. venerealis, and C. fetus subsp. testudinum. Campylobacter fetus fetus affects a broad range of hosts and induces abortion in sheep and cows. Campylobacter fetus venerealis is restricted to cattle and causes the endemic disease bovine genital campylobacteriosis, which triggers reproductive problems and is responsible for major economic losses. Campylobacter fetus testudinum has been proposed recently based on genetically divergent strains isolated from reptiles and humans. Both C. fetus fetus and C. fetus testudinum are opportunistic pathogens for immune-compromised humans. Biochemical tests remain as the gold standard for identifying C. fetus but the fastidious growing requirements and the lack of reliability and reproducibility of some biochemical tests motivated the development of molecular diagnostic tools. These methods have been successfully tested on bovine isolates but fail to detect some genetically divergent strains isolated from other hosts. The aim of the present study was to develop a highly specific molecular assay to identify and quantify C. fetus strains. Results: We developed a highly sensitive real-time PCR assay that targets a unique region of the 16S rRNA gene. This assay successfully detected all C. fetus strains, including those that were negative for the cstA gene-based assay used as a standard for molecular C. fetus identification. The assay showed high specificity and absence of cross-reactivity with other bacterial species. The analytical testing of the assay was determined using a standard curve. The assay demonstrated a wide dynamic range between 102 and 107 genome copies per reaction, and a good reproducibility with small intra- and inter-assay variability. Conclusions: The possibility to characterize samples in a rapid, sensitive and reproducible way makes this assay a good option to establish a new standard in molecular identification and quantification of C. fetus species.
EEA Balcarce
Fil: Iraola, Gregorio. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay. Institut Pasteur Montevideo. Unidad de Bioinformática; Uruguay
Fil: Pérez, Ruben. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay
Fil: Betancor, Laura. Universidad de la República. Facultad de Medicina. Instituto de Higiene. Departamento de Bacteriología y Virología; Uruguay
Fil: Marandino, Ana. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay
Fil: Morsella, Claudia Graciela. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina
Fil: Mendez, Maria Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina
Fil: Paolicchi, Fernando. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina
Fil: Piccirillo, Alessandra. Università degli Studi di Padova. Dipartimento di Biomedicina Comparata e Alimentazione; Italia
Fil: Tomás, Gonzalo. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay
Fil: Velilla, Alejandra Vanesa. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina
Fil: Calleros, Lucía. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay
Fuente
BMC Veterinary Research 12 : 286 (2016)
Materia
Campylobacter fetus
PCR
Genética
Ribosomas
Experimentación en Laboratorio
Genetics
Ribosomes
Laboratory Experimentation
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:20.500.12123/4560
Acceder
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oai_identifier_str oai:localhost:20.500.12123/4560
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network_name_str INTA Digital (INTA)
spelling A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequencesIraola, GregorioPérez, RubenBetancor, LauraMarandino, AnaMorsella, Claudia GracielaMendez, Maria AlejandraPaolicchi, FernandoPiccirillo, AlessandraTomás, GonzaloVelilla, Alejandra VanesaCalleros, LucíaCampylobacter fetusPCRGenéticaRibosomasExperimentación en LaboratorioGeneticsRibosomesLaboratory ExperimentationReacción en Cadena de la PolimerasaBackground: Campylobacter fetus is a pathogen of major concern for animal and human health. The species shows a great intraspecific variation, with three subspecies: C. fetus subsp. fetus, C. fetus subsp. venerealis, and C. fetus subsp. testudinum. Campylobacter fetus fetus affects a broad range of hosts and induces abortion in sheep and cows. Campylobacter fetus venerealis is restricted to cattle and causes the endemic disease bovine genital campylobacteriosis, which triggers reproductive problems and is responsible for major economic losses. Campylobacter fetus testudinum has been proposed recently based on genetically divergent strains isolated from reptiles and humans. Both C. fetus fetus and C. fetus testudinum are opportunistic pathogens for immune-compromised humans. Biochemical tests remain as the gold standard for identifying C. fetus but the fastidious growing requirements and the lack of reliability and reproducibility of some biochemical tests motivated the development of molecular diagnostic tools. These methods have been successfully tested on bovine isolates but fail to detect some genetically divergent strains isolated from other hosts. The aim of the present study was to develop a highly specific molecular assay to identify and quantify C. fetus strains. Results: We developed a highly sensitive real-time PCR assay that targets a unique region of the 16S rRNA gene. This assay successfully detected all C. fetus strains, including those that were negative for the cstA gene-based assay used as a standard for molecular C. fetus identification. The assay showed high specificity and absence of cross-reactivity with other bacterial species. The analytical testing of the assay was determined using a standard curve. The assay demonstrated a wide dynamic range between 102 and 107 genome copies per reaction, and a good reproducibility with small intra- and inter-assay variability. Conclusions: The possibility to characterize samples in a rapid, sensitive and reproducible way makes this assay a good option to establish a new standard in molecular identification and quantification of C. fetus species.EEA BalcarceFil: Iraola, Gregorio. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay. Institut Pasteur Montevideo. Unidad de Bioinformática; UruguayFil: Pérez, Ruben. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; UruguayFil: Betancor, Laura. Universidad de la República. Facultad de Medicina. Instituto de Higiene. Departamento de Bacteriología y Virología; UruguayFil: Marandino, Ana. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; UruguayFil: Morsella, Claudia Graciela. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; ArgentinaFil: Mendez, Maria Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; ArgentinaFil: Paolicchi, Fernando. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; ArgentinaFil: Piccirillo, Alessandra. Università degli Studi di Padova. Dipartimento di Biomedicina Comparata e Alimentazione; ItaliaFil: Tomás, Gonzalo. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; UruguayFil: Velilla, Alejandra Vanesa. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; ArgentinaFil: Calleros, Lucía. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; UruguayBMC2019-03-08T14:37:59Z2019-03-08T14:37:59Z2016-12info: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/s12917-016-0913-3http://hdl.handle.net/20.500.12123/45601746-6148https://doi.org/10.1186/s12917-016-0913-3BMC Veterinary Research 12 : 286 (2016)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)2025-09-29T13:44:35Zoai:localhost:20.500.12123/4560instacron: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 13:44:35.84INTA Digital (INTA) - Instituto Nacional de Tecnología Agropecuariafalse
dc.title.none.fl_str_mv A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences
title A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences
spellingShingle A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences
Iraola, Gregorio
Campylobacter fetus
PCR
Genética
Ribosomas
Experimentación en Laboratorio
Genetics
Ribosomes
Laboratory Experimentation
Reacción en Cadena de la Polimerasa
title_short A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences
title_full A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences
title_fullStr A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences
title_full_unstemmed A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences
title_sort A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences
dc.creator.none.fl_str_mv Iraola, Gregorio
Pérez, Ruben
Betancor, Laura
Marandino, Ana
Morsella, Claudia Graciela
Mendez, Maria Alejandra
Paolicchi, Fernando
Piccirillo, Alessandra
Tomás, Gonzalo
Velilla, Alejandra Vanesa
Calleros, Lucía
author Iraola, Gregorio
author_facet Iraola, Gregorio
Pérez, Ruben
Betancor, Laura
Marandino, Ana
Morsella, Claudia Graciela
Mendez, Maria Alejandra
Paolicchi, Fernando
Piccirillo, Alessandra
Tomás, Gonzalo
Velilla, Alejandra Vanesa
Calleros, Lucía
author_role author
author2 Pérez, Ruben
Betancor, Laura
Marandino, Ana
Morsella, Claudia Graciela
Mendez, Maria Alejandra
Paolicchi, Fernando
Piccirillo, Alessandra
Tomás, Gonzalo
Velilla, Alejandra Vanesa
Calleros, Lucía
author2_role author
author
author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Campylobacter fetus
PCR
Genética
Ribosomas
Experimentación en Laboratorio
Genetics
Ribosomes
Laboratory Experimentation
Reacción en Cadena de la Polimerasa
topic Campylobacter fetus
PCR
Genética
Ribosomas
Experimentación en Laboratorio
Genetics
Ribosomes
Laboratory Experimentation
Reacción en Cadena de la Polimerasa
dc.description.none.fl_txt_mv Background: Campylobacter fetus is a pathogen of major concern for animal and human health. The species shows a great intraspecific variation, with three subspecies: C. fetus subsp. fetus, C. fetus subsp. venerealis, and C. fetus subsp. testudinum. Campylobacter fetus fetus affects a broad range of hosts and induces abortion in sheep and cows. Campylobacter fetus venerealis is restricted to cattle and causes the endemic disease bovine genital campylobacteriosis, which triggers reproductive problems and is responsible for major economic losses. Campylobacter fetus testudinum has been proposed recently based on genetically divergent strains isolated from reptiles and humans. Both C. fetus fetus and C. fetus testudinum are opportunistic pathogens for immune-compromised humans. Biochemical tests remain as the gold standard for identifying C. fetus but the fastidious growing requirements and the lack of reliability and reproducibility of some biochemical tests motivated the development of molecular diagnostic tools. These methods have been successfully tested on bovine isolates but fail to detect some genetically divergent strains isolated from other hosts. The aim of the present study was to develop a highly specific molecular assay to identify and quantify C. fetus strains. Results: We developed a highly sensitive real-time PCR assay that targets a unique region of the 16S rRNA gene. This assay successfully detected all C. fetus strains, including those that were negative for the cstA gene-based assay used as a standard for molecular C. fetus identification. The assay showed high specificity and absence of cross-reactivity with other bacterial species. The analytical testing of the assay was determined using a standard curve. The assay demonstrated a wide dynamic range between 102 and 107 genome copies per reaction, and a good reproducibility with small intra- and inter-assay variability. Conclusions: The possibility to characterize samples in a rapid, sensitive and reproducible way makes this assay a good option to establish a new standard in molecular identification and quantification of C. fetus species.
EEA Balcarce
Fil: Iraola, Gregorio. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay. Institut Pasteur Montevideo. Unidad de Bioinformática; Uruguay
Fil: Pérez, Ruben. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay
Fil: Betancor, Laura. Universidad de la República. Facultad de Medicina. Instituto de Higiene. Departamento de Bacteriología y Virología; Uruguay
Fil: Marandino, Ana. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay
Fil: Morsella, Claudia Graciela. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina
Fil: Mendez, Maria Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina
Fil: Paolicchi, Fernando. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina
Fil: Piccirillo, Alessandra. Università degli Studi di Padova. Dipartimento di Biomedicina Comparata e Alimentazione; Italia
Fil: Tomás, Gonzalo. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay
Fil: Velilla, Alejandra Vanesa. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce. Laboratorio de Bacteriología. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina
Fil: Calleros, Lucía. Universidad de la República. Facultad de Ciencias. Sección Genética Evolutiva; Uruguay
description Background: Campylobacter fetus is a pathogen of major concern for animal and human health. The species shows a great intraspecific variation, with three subspecies: C. fetus subsp. fetus, C. fetus subsp. venerealis, and C. fetus subsp. testudinum. Campylobacter fetus fetus affects a broad range of hosts and induces abortion in sheep and cows. Campylobacter fetus venerealis is restricted to cattle and causes the endemic disease bovine genital campylobacteriosis, which triggers reproductive problems and is responsible for major economic losses. Campylobacter fetus testudinum has been proposed recently based on genetically divergent strains isolated from reptiles and humans. Both C. fetus fetus and C. fetus testudinum are opportunistic pathogens for immune-compromised humans. Biochemical tests remain as the gold standard for identifying C. fetus but the fastidious growing requirements and the lack of reliability and reproducibility of some biochemical tests motivated the development of molecular diagnostic tools. These methods have been successfully tested on bovine isolates but fail to detect some genetically divergent strains isolated from other hosts. The aim of the present study was to develop a highly specific molecular assay to identify and quantify C. fetus strains. Results: We developed a highly sensitive real-time PCR assay that targets a unique region of the 16S rRNA gene. This assay successfully detected all C. fetus strains, including those that were negative for the cstA gene-based assay used as a standard for molecular C. fetus identification. The assay showed high specificity and absence of cross-reactivity with other bacterial species. The analytical testing of the assay was determined using a standard curve. The assay demonstrated a wide dynamic range between 102 and 107 genome copies per reaction, and a good reproducibility with small intra- and inter-assay variability. Conclusions: The possibility to characterize samples in a rapid, sensitive and reproducible way makes this assay a good option to establish a new standard in molecular identification and quantification of C. fetus species.
publishDate 2016
dc.date.none.fl_str_mv 2016-12
2019-03-08T14:37:59Z
2019-03-08T14:37:59Z
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/s12917-016-0913-3
http://hdl.handle.net/20.500.12123/4560
1746-6148
https://doi.org/10.1186/s12917-016-0913-3
url https://bmcvetres.biomedcentral.com/articles/10.1186/s12917-016-0913-3
http://hdl.handle.net/20.500.12123/4560
https://doi.org/10.1186/s12917-016-0913-3
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
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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 12 : 286 (2016)
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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