Development of a Rapid Real-Time PCR Method as a Tool To Quantify Viable Photobacterium phosphoreum Bacteria in Salmon (Salmo salar) Steaks
International audience A specific real-time PCR quantification method combined with a propidium monoazide sample treatment step was developed to determine quantitatively the viable population of the Photobacterium phosphoreum species group in raw modified-atmosphere-packed salmon. Primers were desig...
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Online Access: | https://hal.inrae.fr/hal-02649129 https://doi.org/10.1128/AEM.03677-12 |
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ftunivnantes:oai:HAL:hal-02649129v1 2023-05-15T18:09:58+02:00 Development of a Rapid Real-Time PCR Method as a Tool To Quantify Viable Photobacterium phosphoreum Bacteria in Salmon (Salmo salar) Steaks Mace, Sabrina, Mamlouk, Kelthoum Chipchakova, Stoyka Prévost, Herve, Joffraud, Jean-Jacques Dalgaard, Paw Pilet, Marie-France, Dousset, Xavier SECurité des ALIments et Microbiologie Institut National de la Recherche Agronomique (INRA)-École nationale d'ingénieurs des techniques des industries agricoles et alimentaires (ENITIAA)-École nationale vétérinaire, agroalimentaire et de l'alimentation Nantes-Atlantique (ONIRIS) LUNAM Université Nantes Angers Le Mans Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER) Institut National de la Recherche Agronomique (INRA) Danmarks Tekniske Universitet = Technical University of Denmark (DTU) Bretagne region; Pays de la Loire region; Oniris (Nantes) 2013 https://hal.inrae.fr/hal-02649129 https://doi.org/10.1128/AEM.03677-12 en eng HAL CCSD American Society for Microbiology info:eu-repo/semantics/altIdentifier/doi/10.1128/AEM.03677-12 hal-02649129 https://hal.inrae.fr/hal-02649129 doi:10.1128/AEM.03677-12 PRODINRA: 209075 PUBMEDCENTRAL: PMC3623209 WOS: 000316956200015 ISSN: 0099-2240 EISSN: 1098-5336 Applied and Environmental Microbiology https://hal.inrae.fr/hal-02649129 Applied and Environmental Microbiology, American Society for Microbiology, 2013, 79 (8), pp.2612 - 2619. ⟨10.1128/AEM.03677-12⟩ PATHOGENIC VIBRIO-PARAHAEMOLYTICUS POLYMERASE-CHAIN-REACTION PROPIDIUM MONOAZIDE FISH PRODUCTS BROCHOTHRIX-THERMOSPHACTA LISTERIA-MONOCYTOGENES QUANTITATIVE DETECTION MODIFIED ATMOSPHERES MICROBIAL SPOILAGE RISK-ASSESSMENT [SDV]Life Sciences [q-bio] info:eu-repo/semantics/article Journal articles 2013 ftunivnantes https://doi.org/10.1128/AEM.03677-12 2022-08-10T03:58:10Z International audience A specific real-time PCR quantification method combined with a propidium monoazide sample treatment step was developed to determine quantitatively the viable population of the Photobacterium phosphoreum species group in raw modified-atmosphere-packed salmon. Primers were designed to amplify a 350-bp fragment of the gyrase subunit B gene (gyrB) of P. phosphoreum. The specificity of the two primers was demonstrated by using purified DNA from 81 strains of 52 different bacterial species. When these primers were used for real-time PCR in pure culture, a good correlation (R-2 of 0.99) was obtained between this method and conventional enumeration on marine agar (MA). Quantification was linear over 5 log units as confirmed by using inoculated salmon samples. On naturally contaminated fresh salmon, the new real-time PCR method performed successfully with a quantification limit of 3 log CFU/g. A correlation coefficient (R-2) of 0.963 was obtained between the PCR method and classic enumeration on MA, followed by identification of colonies (290 isolates identified by real-time PCR or by 16S rRNA gene sequencing). A good correlation with an R-2 of 0.940 was found between the new PCR method and an available specific conductance method for P. phosphoreum. This study presents a rapid tool for producing reliable quantitative data on viable P. phosphoreum bacteria in fresh salmon in 6 h. This new culture-independent method will be valuable for future fish inspection, the assessment of raw material quality in fish processing plants, and studies on the ecology of this important specific spoilage microorganism. Article in Journal/Newspaper Salmo salar Université de Nantes: HAL-UNIV-NANTES Applied and Environmental Microbiology 79 8 2612 2619 |
institution |
Open Polar |
collection |
Université de Nantes: HAL-UNIV-NANTES |
op_collection_id |
ftunivnantes |
language |
English |
topic |
PATHOGENIC VIBRIO-PARAHAEMOLYTICUS POLYMERASE-CHAIN-REACTION PROPIDIUM MONOAZIDE FISH PRODUCTS BROCHOTHRIX-THERMOSPHACTA LISTERIA-MONOCYTOGENES QUANTITATIVE DETECTION MODIFIED ATMOSPHERES MICROBIAL SPOILAGE RISK-ASSESSMENT [SDV]Life Sciences [q-bio] |
spellingShingle |
PATHOGENIC VIBRIO-PARAHAEMOLYTICUS POLYMERASE-CHAIN-REACTION PROPIDIUM MONOAZIDE FISH PRODUCTS BROCHOTHRIX-THERMOSPHACTA LISTERIA-MONOCYTOGENES QUANTITATIVE DETECTION MODIFIED ATMOSPHERES MICROBIAL SPOILAGE RISK-ASSESSMENT [SDV]Life Sciences [q-bio] Mace, Sabrina, Mamlouk, Kelthoum Chipchakova, Stoyka Prévost, Herve, Joffraud, Jean-Jacques Dalgaard, Paw Pilet, Marie-France, Dousset, Xavier Development of a Rapid Real-Time PCR Method as a Tool To Quantify Viable Photobacterium phosphoreum Bacteria in Salmon (Salmo salar) Steaks |
topic_facet |
PATHOGENIC VIBRIO-PARAHAEMOLYTICUS POLYMERASE-CHAIN-REACTION PROPIDIUM MONOAZIDE FISH PRODUCTS BROCHOTHRIX-THERMOSPHACTA LISTERIA-MONOCYTOGENES QUANTITATIVE DETECTION MODIFIED ATMOSPHERES MICROBIAL SPOILAGE RISK-ASSESSMENT [SDV]Life Sciences [q-bio] |
description |
International audience A specific real-time PCR quantification method combined with a propidium monoazide sample treatment step was developed to determine quantitatively the viable population of the Photobacterium phosphoreum species group in raw modified-atmosphere-packed salmon. Primers were designed to amplify a 350-bp fragment of the gyrase subunit B gene (gyrB) of P. phosphoreum. The specificity of the two primers was demonstrated by using purified DNA from 81 strains of 52 different bacterial species. When these primers were used for real-time PCR in pure culture, a good correlation (R-2 of 0.99) was obtained between this method and conventional enumeration on marine agar (MA). Quantification was linear over 5 log units as confirmed by using inoculated salmon samples. On naturally contaminated fresh salmon, the new real-time PCR method performed successfully with a quantification limit of 3 log CFU/g. A correlation coefficient (R-2) of 0.963 was obtained between the PCR method and classic enumeration on MA, followed by identification of colonies (290 isolates identified by real-time PCR or by 16S rRNA gene sequencing). A good correlation with an R-2 of 0.940 was found between the new PCR method and an available specific conductance method for P. phosphoreum. This study presents a rapid tool for producing reliable quantitative data on viable P. phosphoreum bacteria in fresh salmon in 6 h. This new culture-independent method will be valuable for future fish inspection, the assessment of raw material quality in fish processing plants, and studies on the ecology of this important specific spoilage microorganism. |
author2 |
SECurité des ALIments et Microbiologie Institut National de la Recherche Agronomique (INRA)-École nationale d'ingénieurs des techniques des industries agricoles et alimentaires (ENITIAA)-École nationale vétérinaire, agroalimentaire et de l'alimentation Nantes-Atlantique (ONIRIS) LUNAM Université Nantes Angers Le Mans Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER) Institut National de la Recherche Agronomique (INRA) Danmarks Tekniske Universitet = Technical University of Denmark (DTU) Bretagne region; Pays de la Loire region; Oniris (Nantes) |
format |
Article in Journal/Newspaper |
author |
Mace, Sabrina, Mamlouk, Kelthoum Chipchakova, Stoyka Prévost, Herve, Joffraud, Jean-Jacques Dalgaard, Paw Pilet, Marie-France, Dousset, Xavier |
author_facet |
Mace, Sabrina, Mamlouk, Kelthoum Chipchakova, Stoyka Prévost, Herve, Joffraud, Jean-Jacques Dalgaard, Paw Pilet, Marie-France, Dousset, Xavier |
author_sort |
Mace, Sabrina, |
title |
Development of a Rapid Real-Time PCR Method as a Tool To Quantify Viable Photobacterium phosphoreum Bacteria in Salmon (Salmo salar) Steaks |
title_short |
Development of a Rapid Real-Time PCR Method as a Tool To Quantify Viable Photobacterium phosphoreum Bacteria in Salmon (Salmo salar) Steaks |
title_full |
Development of a Rapid Real-Time PCR Method as a Tool To Quantify Viable Photobacterium phosphoreum Bacteria in Salmon (Salmo salar) Steaks |
title_fullStr |
Development of a Rapid Real-Time PCR Method as a Tool To Quantify Viable Photobacterium phosphoreum Bacteria in Salmon (Salmo salar) Steaks |
title_full_unstemmed |
Development of a Rapid Real-Time PCR Method as a Tool To Quantify Viable Photobacterium phosphoreum Bacteria in Salmon (Salmo salar) Steaks |
title_sort |
development of a rapid real-time pcr method as a tool to quantify viable photobacterium phosphoreum bacteria in salmon (salmo salar) steaks |
publisher |
HAL CCSD |
publishDate |
2013 |
url |
https://hal.inrae.fr/hal-02649129 https://doi.org/10.1128/AEM.03677-12 |
genre |
Salmo salar |
genre_facet |
Salmo salar |
op_source |
ISSN: 0099-2240 EISSN: 1098-5336 Applied and Environmental Microbiology https://hal.inrae.fr/hal-02649129 Applied and Environmental Microbiology, American Society for Microbiology, 2013, 79 (8), pp.2612 - 2619. ⟨10.1128/AEM.03677-12⟩ |
op_relation |
info:eu-repo/semantics/altIdentifier/doi/10.1128/AEM.03677-12 hal-02649129 https://hal.inrae.fr/hal-02649129 doi:10.1128/AEM.03677-12 PRODINRA: 209075 PUBMEDCENTRAL: PMC3623209 WOS: 000316956200015 |
op_doi |
https://doi.org/10.1128/AEM.03677-12 |
container_title |
Applied and Environmental Microbiology |
container_volume |
79 |
container_issue |
8 |
container_start_page |
2612 |
op_container_end_page |
2619 |
_version_ |
1766182671211298816 |