Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat
Autor(a) principal: | |
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Data de Publicação: | 2023 |
Outros Autores: | , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | https://hdl.handle.net/1822/85560 |
Resumo: | Salmonella Enteritidis is the main serotype responsible for human salmonellosis in the European Union. One of the main sources of Salmonella spp. in the food chain are poultry products, such as eggs or chicken meat. In recent years, molecular methods have become an alternative to culture dependent methods for the rapid screening of Salmonella spp. In this work, the strain S. Enteritidis S1400, and previously isolated and characterized bacteriophage PVP-SE2, were used to develop and evaluate a same-day detection method combining Phage Amplification and Loop-mediated isothermal amplification (PA-LAMP) to specifically detect viable S. Enteritidis in chicken breast. This method is based on the detection of the phage DNA rather than bacterial DNA. The virus is added to the sample during pre-enrichment in buffered peptone water, where it replicates in the presence of viable S. Enteritidis. The detection of phage DNA allows, on the one hand to detect viable bacteria, since viruses only replicate in them, and on the other hand to increase the sensitivity of the method since for each infected S. Enteritidis cell, hundreds of new viruses are produced. Two different PA-LAMP detection strategies were evaluated, a real time fluorescence and a naked-eye detection. The present method could down to 0.2 fg/L of pure phage DNA and a concentration of viral particles of 2.2 log PFU/mL. After a short Salmonella recovery step of 3 h and a co-culture of 4 h of the samples with phage particles, both real-time fluorescence and naked-eye method showed a LoD95 of 6.6 CFU/25 g and a LoD50 of 1.5/25 g in spiked chicken breast samples. The entire detection process, including DNA extraction and LAMP analysis, can be completed in around 8 h. In the current proof-of-concept, the novel PA-LAMP obtained comparable results to those of the reference method ISO 6579, to detect Salmonella Enteritidis in poultry meat. |
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Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meatPhage amplificationLAMPEnteritidisDetectionChickenPoultry meatVisual detectionSalmonella EnteritidisSalmonella Enteritidis is the main serotype responsible for human salmonellosis in the European Union. One of the main sources of Salmonella spp. in the food chain are poultry products, such as eggs or chicken meat. In recent years, molecular methods have become an alternative to culture dependent methods for the rapid screening of Salmonella spp. In this work, the strain S. Enteritidis S1400, and previously isolated and characterized bacteriophage PVP-SE2, were used to develop and evaluate a same-day detection method combining Phage Amplification and Loop-mediated isothermal amplification (PA-LAMP) to specifically detect viable S. Enteritidis in chicken breast. This method is based on the detection of the phage DNA rather than bacterial DNA. The virus is added to the sample during pre-enrichment in buffered peptone water, where it replicates in the presence of viable S. Enteritidis. The detection of phage DNA allows, on the one hand to detect viable bacteria, since viruses only replicate in them, and on the other hand to increase the sensitivity of the method since for each infected S. Enteritidis cell, hundreds of new viruses are produced. Two different PA-LAMP detection strategies were evaluated, a real time fluorescence and a naked-eye detection. The present method could down to 0.2 fg/L of pure phage DNA and a concentration of viral particles of 2.2 log PFU/mL. After a short Salmonella recovery step of 3 h and a co-culture of 4 h of the samples with phage particles, both real-time fluorescence and naked-eye method showed a LoD95 of 6.6 CFU/25 g and a LoD50 of 1.5/25 g in spiked chicken breast samples. The entire detection process, including DNA extraction and LAMP analysis, can be completed in around 8 h. In the current proof-of-concept, the novel PA-LAMP obtained comparable results to those of the reference method ISO 6579, to detect Salmonella Enteritidis in poultry meat.Dr. Alexandre Lamas was funded by a postdoctoral fellowship from Xunta de Galicia (Axudas de apoio a étapa de formación posdoutoral IN606B (Modalidade A)). Dr. Alejandro Garrido-Maestu acknowledges funding from the Fundação para a Ciência e Tecnologia through the Scientific Employment Stimulus Program (2021.02810.CEECIND).info:eu-repo/semantics/publishedVersionElsevierUniversidade do MinhoLamas, A.Santos, Sílvio Roberto BrancoMarta PradoGarrido-Maestu, A.2023-102023-10-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttps://hdl.handle.net/1822/85560engLamas, A., Santos, S. B., Prado, M., & Garrido-Maestu, A. (2023, October). Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat. Food Microbiology. Elsevier BV. http://doi.org/10.1016/j.fm.2023.1043410740-002010.1016/j.fm.2023.10434137567642104341https://www.sciencedirect.com/science/article/pii/S0740002023001284info:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2023-09-30T01:25:43Zoai:repositorium.sdum.uminho.pt:1822/85560Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-19T18:46:24.394443Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
dc.title.none.fl_str_mv |
Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat |
title |
Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat |
spellingShingle |
Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat Lamas, A. Phage amplification LAMP Enteritidis Detection Chicken Poultry meat Visual detection Salmonella Enteritidis |
title_short |
Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat |
title_full |
Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat |
title_fullStr |
Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat |
title_full_unstemmed |
Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat |
title_sort |
Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat |
author |
Lamas, A. |
author_facet |
Lamas, A. Santos, Sílvio Roberto Branco Marta Prado Garrido-Maestu, A. |
author_role |
author |
author2 |
Santos, Sílvio Roberto Branco Marta Prado Garrido-Maestu, A. |
author2_role |
author author author |
dc.contributor.none.fl_str_mv |
Universidade do Minho |
dc.contributor.author.fl_str_mv |
Lamas, A. Santos, Sílvio Roberto Branco Marta Prado Garrido-Maestu, A. |
dc.subject.por.fl_str_mv |
Phage amplification LAMP Enteritidis Detection Chicken Poultry meat Visual detection Salmonella Enteritidis |
topic |
Phage amplification LAMP Enteritidis Detection Chicken Poultry meat Visual detection Salmonella Enteritidis |
description |
Salmonella Enteritidis is the main serotype responsible for human salmonellosis in the European Union. One of the main sources of Salmonella spp. in the food chain are poultry products, such as eggs or chicken meat. In recent years, molecular methods have become an alternative to culture dependent methods for the rapid screening of Salmonella spp. In this work, the strain S. Enteritidis S1400, and previously isolated and characterized bacteriophage PVP-SE2, were used to develop and evaluate a same-day detection method combining Phage Amplification and Loop-mediated isothermal amplification (PA-LAMP) to specifically detect viable S. Enteritidis in chicken breast. This method is based on the detection of the phage DNA rather than bacterial DNA. The virus is added to the sample during pre-enrichment in buffered peptone water, where it replicates in the presence of viable S. Enteritidis. The detection of phage DNA allows, on the one hand to detect viable bacteria, since viruses only replicate in them, and on the other hand to increase the sensitivity of the method since for each infected S. Enteritidis cell, hundreds of new viruses are produced. Two different PA-LAMP detection strategies were evaluated, a real time fluorescence and a naked-eye detection. The present method could down to 0.2 fg/L of pure phage DNA and a concentration of viral particles of 2.2 log PFU/mL. After a short Salmonella recovery step of 3 h and a co-culture of 4 h of the samples with phage particles, both real-time fluorescence and naked-eye method showed a LoD95 of 6.6 CFU/25 g and a LoD50 of 1.5/25 g in spiked chicken breast samples. The entire detection process, including DNA extraction and LAMP analysis, can be completed in around 8 h. In the current proof-of-concept, the novel PA-LAMP obtained comparable results to those of the reference method ISO 6579, to detect Salmonella Enteritidis in poultry meat. |
publishDate |
2023 |
dc.date.none.fl_str_mv |
2023-10 2023-10-01T00:00:00Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
https://hdl.handle.net/1822/85560 |
url |
https://hdl.handle.net/1822/85560 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
Lamas, A., Santos, S. B., Prado, M., & Garrido-Maestu, A. (2023, October). Phage amplification coupled with loop-mediated isothermal amplification (PA-LAMP) for same-day detection of viable Salmonella Enteritidis in raw poultry meat. Food Microbiology. Elsevier BV. http://doi.org/10.1016/j.fm.2023.104341 0740-0020 10.1016/j.fm.2023.104341 37567642 104341 https://www.sciencedirect.com/science/article/pii/S0740002023001284 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.publisher.none.fl_str_mv |
Elsevier |
publisher.none.fl_str_mv |
Elsevier |
dc.source.none.fl_str_mv |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
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