Development of a mreB-targeted real-time PCR method for the quantitative detection of Vibrio harveyi in seawater and biofilm from aquaculture systems
| العنوان: | Development of a mreB-targeted real-time PCR method for the quantitative detection of Vibrio harveyi in seawater and biofilm from aquaculture systems |
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| المؤلفون: | Marie-Agnes Travers, Thierry Grard, Roxane Roquigny, Maryse Bonnin-Jusserand, Julia Mougin, Cédric Le Bris |
| المساهمون: | Transfrontalière BioEcoAgro (Transfrontalière BioEcoAgro), Université d'Artois (UA)-Université de Liège-Université de Picardie Jules Verne (UPJV)-Université du Littoral Côte d'Opale (ULCO)-Université de Lille-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Interactions Hôtes-Pathogènes-Environnements (IHPE), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER)-Université de Perpignan Via Domitia (UPVD), Laboratoire de Génétique et Pathologie des Mollusques Marins, 17390 La Tremblade, France. (LGPMM), Santé, Génétique et Microbiologie des Mollusques (IFREMER SG2M), Institut Français de Recherche pour l'Exploitation de la Mer - Atlantique (IFREMER Atlantique), Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER)-Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER)-Institut Français de Recherche pour l'Exploitation de la Mer - Atlantique (IFREMER Atlantique), Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER)-Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER), Hauts-de-France regional council, ULCO, French government, Ifremer, Region Hauts-de-France, FEAMP as part of the LUVIBAR project PFEA470017FA1000006, Transfrontalière BioEcoAgro - UMR 1158 (BioEcoAgro), Université d'Artois (UA)-Université de Liège-Université de Picardie Jules Verne (UPJV)-Université du Littoral Côte d'Opale (ULCO)-Université de Lille-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-JUNIA (JUNIA), Université catholique de Lille (UCL)-Université catholique de Lille (UCL), Biochimie des Produits Aquatiques (BPA), Université du Littoral Côte d'Opale (ULCO)-Transfrontalière BioEcoAgro - UMR 1158 (BioEcoAgro), Université catholique de Lille (UCL)-Université catholique de Lille (UCL)-Université d'Artois (UA)-Université de Liège-Université de Picardie Jules Verne (UPJV)-Université du Littoral Côte d'Opale (ULCO)-Université de Lille-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-JUNIA (JUNIA), Université de Perpignan Via Domitia (UPVD)-Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Laboratoire de Génétique et Pathologie des Mollusques Marins (LGPMM), Santé, Génétique et Microbiologie des Mollusques (SGMM) |
| المصدر: | Aquaculture Aquaculture, Elsevier, 2020, 525, pp.735337. ⟨10.1016/j.aquaculture.2020.735337⟩ Aquaculture, 2020, 525, pp.735337. ⟨10.1016/j.aquaculture.2020.735337⟩ Aquaculture (0044-8486) (Elsevier BV), 2020-08, Vol. 525, P. 735337 (9p.) |
| بيانات النشر: | HAL CCSD, 2020. |
| سنة النشر: | 2020 |
| مصطلحات موضوعية: | [SDV.SA.ZOO]Life Sciences [q-bio]/Agricultural sciences/Zootechny, Aquaculture, Aquatic Science, Genome, MreB, Microbiology, law.invention, 03 medical and health sciences, chemistry.chemical_compound, law, Seawater, 14. Life underwater, Polymerase chain reaction, 030304 developmental biology, 0303 health sciences, biology, Vibrio harveyi, Biofilm, fungi, 04 agricultural and veterinary sciences, biology.organism_classification, Quantitative detection, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, mreB gene, Real-time polymerase chain reaction, chemistry, 040102 fisheries, SYBR Green I, 0401 agriculture, forestry, and fisheries, bacteria, Bacteria |
| الوصف: | International audience; Vibrio harveyi is a particularly problematic Gram-negative bacterium because it can form biofilms on aquaculture facility surfaces, leading to resistance of bacteria against antibiotics and water sanitizers. A SYBR Green I quantitative real-time PCR method was developed to detect V. harveyi directly from environmental samples, including seawater and biofilm. Specific primers targeting the mreB gene were designed. The exclusivity and inclusivity of the newly designed primers were evaluated using a panel of 85 bacteria: 58 V. harveyi from multiples origins and 27 non-V. harveyi isolates, and compared with two pairs of primers targeting the topA and toxR genes that were designed previously. All sets of primers were able to distinguish V. harveyi from closely related species belonging to the Harveyi clade. However, the mreB primers showed better inclusivity and were thus used to develop the real-time PCR assay. A quantification curve was obtained from pure culture of V. harveyi and exhibited excellent efficacy with detection levels as low as 5 genome copies in the PCR reaction. After selection of the extraction kit allowing the best DNA quantity and purity, validation was performed on both seawater and biofilm samples collected from a fish farm. The presence of inhibitors in the DNA templates was evaluated and a 10-fold dilution of template DNA was recommended in order to avoid their effects. The assay was able to detect V. harveyi from environmental samples, confirming the validity of the method. This real-time PCR method will help to evaluate the dynamics of V. harveyi in aquaculture facilities. Suitable prophylactic control measures could be designed using this method, instead of the use of curative methods such as antibiotics |
| وصف الملف: | application/pdf |
| اللغة: | English |
| تدمد: | 0044-8486 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::1675173137a8c2a72498426371cbd20d https://hal.archives-ouvertes.fr/hal-02864334/file/Mougin-2020-Aquaculture-Developpement-MANUSCRIT.pdf |
| حقوق: | OPEN |
| رقم الأكسشن: | edsair.doi.dedup.....1675173137a8c2a72498426371cbd20d |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 00448486 |
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