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
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 experime...
| Autores principales: | , , , , , , |
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| Formato: | info:eu-repo/semantics/article |
| Lenguaje: | Inglés |
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2017
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| Acceso en línea: | http://hdl.handle.net/20.500.12123/1742 https://www.sciencedirect.com/science/article/pii/S002364381730155X https://doi.org/10.1016/j.lwt.2017.03.010 |
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| author | Pega, Juan Franco Descalzo, Adriana Maria Rossetti, Luciana Nanni, Mariana Perez, Carolina Daiana Rizzo, Sergio Anibal Diaz, Gabriela Perez |
| author_browse | Descalzo, Adriana Maria Diaz, Gabriela Perez Nanni, Mariana Pega, Juan Franco Perez, Carolina Daiana Rizzo, Sergio Anibal Rossetti, Luciana |
| author_facet | Pega, Juan Franco Descalzo, Adriana Maria Rossetti, Luciana Nanni, Mariana Perez, Carolina Daiana Rizzo, Sergio Anibal Diaz, Gabriela Perez |
| author_sort | Pega, Juan Franco |
| collection | INTA Digital |
| 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. |
| format | info:eu-repo/semantics/article |
| id | INTA1742 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2017 |
| publishDateRange | 2017 |
| publishDateSort | 2017 |
| record_format | dspace |
| spelling | INTA17422018-06-21T12:37:41Z 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 Descalzo, Adriana Maria Rossetti, Luciana Nanni, Mariana Perez, Carolina Daiana Rizzo, Sergio Anibal Diaz, Gabriela Perez Acidos Nucléicos Bacterias Acidolácticas Marcadores Genéticos Productos Lácteos Nucleic Acids Lactic Acid Bacteria Genetic Markers PCR Milk Products Marcadores Moleculares Reacción en Cadena de la Polimerasa 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. Inst. de Tecnol. de los Alimentos- ITA Fil: Pega, Juan Franco. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Descalzo, Adriana Maria. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto Tecnología de Alimentos; Argentina. Instituto Nacional de Tecnología Agropecuaria (INTA).LABINTEX-Centre de Coopération Internationale en Recherche Agronomique pour le Déeveloppement (CIRAD), Déepartement PERSYST, UMR Qualisud; Francia Fil: Rossetti, Luciana. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto Tecnología de Alimentos; Argentina Fil: Perez, Carolina Daiana. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto Tecnología de Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Diaz, Gabriela Perez. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto Tecnología de Alimentos; Argentina Fil: Nanni, Mariana. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto Tecnología de Alimentos; Argentina Fil: Rizzo, Sergio Anibal. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto Tecnología de Alimentos; Argentina 2017-11-10T12:07:47Z 2017-11-10T12:07:47Z 2017-07 info:eu-repo/semantics/article info:ar-repo/semantics/artículo info:eu-repo/semantics/acceptedVersion http://hdl.handle.net/20.500.12123/1742 https://www.sciencedirect.com/science/article/pii/S002364381730155X 0023-6438 https://doi.org/10.1016/j.lwt.2017.03.010 eng info:eu-repo/semantics/restrictedAccess application/pdf LWT - Food science and technology 80 : 416-422. (July 2017) |
| spellingShingle | Acidos Nucléicos Bacterias Acidolácticas Marcadores Genéticos Productos Lácteos Nucleic Acids Lactic Acid Bacteria Genetic Markers PCR Milk Products Marcadores Moleculares Reacción en Cadena de la Polimerasa Pega, Juan Franco Descalzo, Adriana Maria Rossetti, Luciana Nanni, Mariana Perez, Carolina Daiana Rizzo, Sergio Anibal Diaz, Gabriela Perez 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 |
| 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_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_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 |
| topic | Acidos Nucléicos Bacterias Acidolácticas Marcadores Genéticos Productos Lácteos Nucleic Acids Lactic Acid Bacteria Genetic Markers PCR Milk Products Marcadores Moleculares Reacción en Cadena de la Polimerasa |
| url | http://hdl.handle.net/20.500.12123/1742 https://www.sciencedirect.com/science/article/pii/S002364381730155X https://doi.org/10.1016/j.lwt.2017.03.010 |
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