Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity

Autores
Pega, Juan Franco; Rizzo, Sergio Anibal; Rossetti, Luciana; Pérez, Carolina Daiana; Díaz, G.; Descalzo, Adriana Maria; Nanni, Mariana Sandra
Año de publicación
2017
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
In this study, we evaluated whether quantitative PCR (qPCR) and reverse transcription-qPCR (RT-qPCR) results could be affected by extracellular (free) DNA or RNA, respectively, obtained either from isolated bacteria or from fermented dairy foods manufactured with these microorganisms. These experimental set-ups involved two starter lactic acid bacteria (SLAB), Streptococcus thermophilus and Lactococcus lactis, as models across their most common technological environments: yoghurt, milk and cheese. The presence of free DNA modified qPCR results in levels sufficient to bias SLAB determinations, indicating that DNA may be a poor measure of cellular integrity. Furthermore, we showed for the first time that an order of magnitude more of free RNA than of DNA had to be used to modify RT-qPCR results. Importantly, the interference produced by free DNA and RNA was maintained across all the dairy matrices evaluated, independently of the SLAB and of the source of nucleic acids used, in both negative and positive samples. This study suggests that, contrary to DNA molecules, mRNA from dead microorganisms or from other extracellular sources may not be a determinant factor in skewing molecular quantifications. Therefore, our results support the potential use of mRNA as a molecular signature of cell integrity in dairy matrices.
Fil: Pega, Juan Franco. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Rizzo, Sergio Anibal. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina
Fil: Rossetti, Luciana. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina
Fil: Pérez, Carolina Daiana. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Díaz, G.. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina
Fil: Descalzo, Adriana Maria. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Centre de Coopération Internationale en Recherche Agronomique pour le Développerment; Francia
Fil: Nanni, Mariana Sandra. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Materia
Cell Integrity
Dairy Foods
Qpcr
Rt-Qpcr
Slab
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/72847
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/72847
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrityPega, Juan FrancoRizzo, Sergio AnibalRossetti, LucianaPérez, Carolina DaianaDíaz, G.Descalzo, Adriana MariaNanni, Mariana SandraCell IntegrityDairy FoodsQpcrRt-QpcrSlabhttps://purl.org/becyt/ford/2.11https://purl.org/becyt/ford/2In this study, we evaluated whether quantitative PCR (qPCR) and reverse transcription-qPCR (RT-qPCR) results could be affected by extracellular (free) DNA or RNA, respectively, obtained either from isolated bacteria or from fermented dairy foods manufactured with these microorganisms. These experimental set-ups involved two starter lactic acid bacteria (SLAB), Streptococcus thermophilus and Lactococcus lactis, as models across their most common technological environments: yoghurt, milk and cheese. The presence of free DNA modified qPCR results in levels sufficient to bias SLAB determinations, indicating that DNA may be a poor measure of cellular integrity. Furthermore, we showed for the first time that an order of magnitude more of free RNA than of DNA had to be used to modify RT-qPCR results. Importantly, the interference produced by free DNA and RNA was maintained across all the dairy matrices evaluated, independently of the SLAB and of the source of nucleic acids used, in both negative and positive samples. This study suggests that, contrary to DNA molecules, mRNA from dead microorganisms or from other extracellular sources may not be a determinant factor in skewing molecular quantifications. Therefore, our results support the potential use of mRNA as a molecular signature of cell integrity in dairy matrices.Fil: Pega, Juan Franco. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Rizzo, Sergio Anibal. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; ArgentinaFil: Rossetti, Luciana. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; ArgentinaFil: Pérez, Carolina Daiana. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Díaz, G.. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; ArgentinaFil: Descalzo, Adriana Maria. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Centre de Coopération Internationale en Recherche Agronomique pour le Développerment; FranciaFil: Nanni, Mariana Sandra. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaElsevier Science2017-07info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/72847Pega, Juan Franco; Rizzo, Sergio Anibal; Rossetti, Luciana; Pérez, Carolina Daiana; Díaz, G.; et al.; Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity; Elsevier Science; LWT - Food Science and Technology; 80; 7-2017; 416-4220023-6438CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1016/j.lwt.2017.03.010info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S002364381730155Xinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-nd/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:09:31Zoai:ri.conicet.gov.ar:11336/72847instacron: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:34982025-09-29 10:09:31.256CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity
title Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity
spellingShingle Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity
Pega, Juan Franco
Cell Integrity
Dairy Foods
Qpcr
Rt-Qpcr
Slab
title_short Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity
title_full Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity
title_fullStr Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity
title_full_unstemmed Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity
title_sort Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity
dc.creator.none.fl_str_mv Pega, Juan Franco
Rizzo, Sergio Anibal
Rossetti, Luciana
Pérez, Carolina Daiana
Díaz, G.
Descalzo, Adriana Maria
Nanni, Mariana Sandra
author Pega, Juan Franco
author_facet Pega, Juan Franco
Rizzo, Sergio Anibal
Rossetti, Luciana
Pérez, Carolina Daiana
Díaz, G.
Descalzo, Adriana Maria
Nanni, Mariana Sandra
author_role author
author2 Rizzo, Sergio Anibal
Rossetti, Luciana
Pérez, Carolina Daiana
Díaz, G.
Descalzo, Adriana Maria
Nanni, Mariana Sandra
author2_role author
author
author
author
author
author
dc.subject.none.fl_str_mv Cell Integrity
Dairy Foods
Qpcr
Rt-Qpcr
Slab
topic Cell Integrity
Dairy Foods
Qpcr
Rt-Qpcr
Slab
purl_subject.fl_str_mv https://purl.org/becyt/ford/2.11
https://purl.org/becyt/ford/2
dc.description.none.fl_txt_mv In this study, we evaluated whether quantitative PCR (qPCR) and reverse transcription-qPCR (RT-qPCR) results could be affected by extracellular (free) DNA or RNA, respectively, obtained either from isolated bacteria or from fermented dairy foods manufactured with these microorganisms. These experimental set-ups involved two starter lactic acid bacteria (SLAB), Streptococcus thermophilus and Lactococcus lactis, as models across their most common technological environments: yoghurt, milk and cheese. The presence of free DNA modified qPCR results in levels sufficient to bias SLAB determinations, indicating that DNA may be a poor measure of cellular integrity. Furthermore, we showed for the first time that an order of magnitude more of free RNA than of DNA had to be used to modify RT-qPCR results. Importantly, the interference produced by free DNA and RNA was maintained across all the dairy matrices evaluated, independently of the SLAB and of the source of nucleic acids used, in both negative and positive samples. This study suggests that, contrary to DNA molecules, mRNA from dead microorganisms or from other extracellular sources may not be a determinant factor in skewing molecular quantifications. Therefore, our results support the potential use of mRNA as a molecular signature of cell integrity in dairy matrices.
Fil: Pega, Juan Franco. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Rizzo, Sergio Anibal. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina
Fil: Rossetti, Luciana. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina
Fil: Pérez, Carolina Daiana. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Díaz, G.. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina
Fil: Descalzo, Adriana Maria. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Centre de Coopération Internationale en Recherche Agronomique pour le Développerment; Francia
Fil: Nanni, Mariana Sandra. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación de Agroindustria. Instituto de Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
description In this study, we evaluated whether quantitative PCR (qPCR) and reverse transcription-qPCR (RT-qPCR) results could be affected by extracellular (free) DNA or RNA, respectively, obtained either from isolated bacteria or from fermented dairy foods manufactured with these microorganisms. These experimental set-ups involved two starter lactic acid bacteria (SLAB), Streptococcus thermophilus and Lactococcus lactis, as models across their most common technological environments: yoghurt, milk and cheese. The presence of free DNA modified qPCR results in levels sufficient to bias SLAB determinations, indicating that DNA may be a poor measure of cellular integrity. Furthermore, we showed for the first time that an order of magnitude more of free RNA than of DNA had to be used to modify RT-qPCR results. Importantly, the interference produced by free DNA and RNA was maintained across all the dairy matrices evaluated, independently of the SLAB and of the source of nucleic acids used, in both negative and positive samples. This study suggests that, contrary to DNA molecules, mRNA from dead microorganisms or from other extracellular sources may not be a determinant factor in skewing molecular quantifications. Therefore, our results support the potential use of mRNA as a molecular signature of cell integrity in dairy matrices.
publishDate 2017
dc.date.none.fl_str_mv 2017-07
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/11336/72847
Pega, Juan Franco; Rizzo, Sergio Anibal; Rossetti, Luciana; Pérez, Carolina Daiana; Díaz, G.; et al.; Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity; Elsevier Science; LWT - Food Science and Technology; 80; 7-2017; 416-422
0023-6438
CONICET Digital
CONICET
url http://hdl.handle.net/11336/72847
identifier_str_mv Pega, Juan Franco; Rizzo, Sergio Anibal; Rossetti, Luciana; Pérez, Carolina Daiana; Díaz, G.; et al.; Impact of extracellular nucleic acids from lactic acid bacteria on qPCR and RT-qPCR results in dairy matrices: Implications for defining molecular markers of cell integrity; Elsevier Science; LWT - Food Science and Technology; 80; 7-2017; 416-422
0023-6438
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1016/j.lwt.2017.03.010
info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S002364381730155X
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Elsevier Science
publisher.none.fl_str_mv Elsevier Science
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 13.070432

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