Pathogenicity of Vibrio penaeicida or white shrimp Litopenaeus vannamei: a cysteine protease-like exotoxin as a virulence factor Auteur(s) : Aguirre Guzman, G Ascencio, F Saulnier, Denis Éditeur(s) : Inter-Research Résumé : The pathogenicity of Vibrio penaeicida Strains KH-1 and AM101, their culture-free supernatant (CFS), and their protein fraction obtained by 40% of ammonium sulfate precipitation (PFs40) were assessed in experimental challenges against juvenile Litopenaeus vannamei. Live Vibrio cells, CFS, and PFs40 from the AM101 strain produced a significantly higher mortality (p < 0.05) compared to the KH-1 strain. Toxicity and median lethal doses (LD50) of Fast Protein Liquid Chromatography (FPLC) products were evaluated on L. vannamei. The first FPLC fraction sample (A) from PFs40 of the AM101 strain displayed LD50 values of 1.68 and 5.61 mu g protein ind.(-1), respectively. The second FPLC process from Fraction A showed a peak (A1) also with toxic effects to shrimp. PFs40, Fraction A, and Peak At showed a 38.5 kDa molecular band (SDS-PAGE), with activity on a gelatin protease zymogram. The lethal effect of PFs40 and Fraction A was inhibited by Proteinase K, CuCl2, E-64, and heat (60 and 100 degrees C) treatments, but was not inhibited by EDTA-Na-2, aprotinin, and soy trypsin treatments. These results and the zymogram inhibition test suggest the presence of a cysteine protease-like proteinaceous exotoxin as a dominant protease, secreted by V. penaeicida Strain AM101. Diseases of aquatic organisms (0177-5103) (Inter-Research), 2005-11 , Vol. 67 , N. 3 , P. 201-207 Droits : Inter-Research 2005 http://archimer.ifremer.fr/doc/2005/publication-3618.pdf DOI:10.3354/dao067201 http://archimer.ifremer.fr/doc/00000/3618/ | Partager |
Aspects moléculaires et biochimiques des stylicines, peptides multifonctionnels identifiés chez la crevette bleue du Pacifique Litopenaeus stylirostris (Crustacea, Decapoda) Auteur(s) : Rolland, Jean-luc Éditeur(s) : Université Montpellier 2 - Sciences et Techniques du Languedoc Résumé : The work reported here was motivated by the economical importance of the pacific blue shrimp Litopenaeus stylirostris farming where high mortality rates are due to bacterial and viral diseases. It consists in the characterisation of two original peptides, the first members of a new multifunctional family of peptides from peneide shrimps, the stylicines. Those two peptides, named stylicines 1 and 2, are negatively charged (pI < 6.0), and characterised by a proline-rich N-terminal region and a C-terminal region containing 13 cysteine residues. Stylicines are synthesized by heamocytes where they are stored within small cytoplasmic granules. To understand the role of these peptides in the immune response of shrimps to a vibrio infection, their recombinant forms were produced in E. coli BL21 (DE3) plysS, purified and characterised. The two rstylicines display biological anti-proliferative and blood clotting activities. Only rstylicine 1 displays antimicrobial activities: antifungal against Fusarium oxysporum (MIC<2.5μM) and bacteriostatic against Gram (−) bacteria, Vibrio sp. (MIC<80μM). Moreover this peptide displays an LPS-binding activity (dissociation constant (Kd) of 9.6×10−8 M) and agglutinate Vibrio. penaeicida "in vitro". Finally, the presence of sequences coding for modified forms of stylicine 1 in some shrimp’s genome may be in relation with their lower ability to survive infections. Les travaux présentés dans ce mémoire ont été motivés par l’importance économique de l’élevage de la crevette bleue du pacifique Litopenaeus stylirostris dont les fortes mortalités sont principalement dues au développement de maladies bactériennes et virales. Ils ont consisté en la caractérisation des deux premiers membres d’une famille originale de peptides multifonctionnels présents chez les crevettes pénéides, les stylicines. Ces peptides, nommés stylicines 1 et 2, sont des peptides anioniques (pI < 6.0), formés d’une région amino-terminale riche en résidus de type proline et d’une région carboxyterminale riche de treize résidus cystéines. Ces molécules sont synthétisées et stockées dans de petits granules présents dans le cytoplasme des hémocytes. Pour mieux appréhender leurs rôles dans la réponse immunitaire des crevettes à une infection par des Vibrio, leurs formes recombinantes ont été produites dans E. coli BL21 (DE3) plysS, purifiées et caractérisées. Les deux rstylicines présentent des activités antiproliférative et anticoagulante. Seule la rstylicine1 présente des activités antimicrobiennes : antifongique sur Fusarium oxysporum (CMI<2.5 μM), et antibactérienne (bactériostatique) sur Vibrio sp (CMI<80 μM). Ce peptide est également capable de se lier aux LPS des bactéries à Gram (-) (Kd= 9.6x10-8 M) et d’agglutiner V. penaeicida "in vitro". Enfin, l’existence de gènes codant des formes modifiées de la stylicine1, chez certaines crevettes, pourrait être en relation avec une diminution de la résistante des individus aux infections. Droits : 2010 The Author, Univ Montpellier 2 http://archimer.ifremer.fr/doc/00024/13558/10612.pdf http://archimer.ifremer.fr/doc/00024/13558/ | Partager |
Toxic factors of Vibrio strains pathogenic to shrimp Auteur(s) : Goarant, Cyrille Herlin, Jose Brizard, Raphael Marteau, Anne-laure Martin, C Martin, B Éditeur(s) : Inter-research Résumé : Vibriosis is a major disease problem in shrimp aquaculture. 'Syndrome 93' is a seasonal juvenile vibriosis caused by Vibrio penaeicida which affects Litopenaeus stylirostris in grow-out ponds in New Caledonia. This study assessed the toxic activities of extracellular products (ECPs) from V: penaeicida, V. alginolyticus and V, nigripulchritudo using in vivo injections in healthy juvenile L. stylirostris (= Penaeus stylirostris) and in vitro assays on shrimp primary cell cultures and the fish cell line epithelioma papulosum cyprini (EPC). Toxic effects of ECPs were demonstrated for all pathogenic Vibrio strains tested both in vivo and in vitro, but for shrimp only; no effect was observed on the fish cell line. ECP toxicity for New Caledonian V. penaeicida was found only after cultivation at low temperature (20 degrees C) and not at higher temperature (30 degrees C). This points to the fact that 'Syndrome 93' episodes are triggered by temperature drops. The assays used here demonstrate the usefulness of primary shrimp cell cultures to study virulence mechanisms of shrimp pathogenic bacteria. Diseases Of Aquatic Organisms (0177-5103) (Inter-research), 2000-03 , Vol. 40 , N. 2 , P. 101-107 Droits : info:eu-repo/semantics/openAccess http://archimer.ifremer.fr/doc/00000/10543/7600.pdf DOI:10.3354/dao040101 http://archimer.ifremer.fr/doc/00000/10543/ | Partager |
Rapid and sensitive PCR detection of Vibrio penaeicida, the putative etiological agent of syndrome 93 in New Caledonia Auteur(s) : Saulnier, Denis Avarre, Jean-christophe Le Moullac, Gilles Ansquer, Dominique Levy, Peva Vonau, Vincent Éditeur(s) : Inter-Research Résumé : Experimental infections of Penaeus (Litopenaeus) stylirostris were performed with a Vibrio penaeicida strain (AM101) isolated in New Caledonia from Syndrome 93 diseased shrimp. Cumulative mortalities resulting from intramuscular injection or immersion of shrimp in bacterial suspensions demonstrated high virulence for this bacterial strain and suggested that V. penaeicida could be the etiological agent of Syndrome 93. The median lethal dose (LD50) for AM101 was 1.3 x 104 CFU (colony forming units) ml-1 by immersion and less than 5 CFU shrimp-1 by intramuscular challenge, with mortality outbreaks at 48 and 22 h after challenge, respectively. A polymerase chain reaction (PCR) detection assay using a primer set designed from the 16S ribosomal RNA gene of V. penaeicida was developed. It gave an expected amplicon of approximately 310 bp in ethidium bromide-stained agarose gels. The specificity of these primers was assessed with different Vibrio species. Furthermore, DNA extracted by the ChelexTM method could be used to detect fewer than 20 cultured Vibrio cells in seawater or shrimp hemolymph by this assay. It appears to be a reliable screening method for detecting V. penaeicida in shrimp and from the aquatic environment. Diseases of aquatic organisms (0177-5103) (Inter-Research), 2000-03 , Vol. 49 , N. 2 , P. 109-115 Droits : Inter-Research 2000 http://archimer.ifremer.fr/doc/2000/publication-4436.pdf DOI:10.3354/dao040109 http://archimer.ifremer.fr/doc/00000/4436/ | Partager |
Résistance de la crevette Litopenaeus stylirostris à la bactérie pathogène Vibrio penaeicida : Physiologie, immunologie et pathologie comparées d’une population sélectionnée sur un critère de survie aux épisodes de mortalité et d’une population témoin non sélectionnée. Auteur(s) : De Decker, Sophie Résumé : The New-Caledonian shrimp industry is based on the controlled reproduction of the shrimp Litopenaeus stylirostris, a species which was introduced in the 80s. The major difficulty to which the industry has been faced for 10 years is the occurrence of the “Syndrome 93”, which corresponds to mortality phases when the temperature falls down in April-May-June. This mortality is associated to the pathogenic bacteria Vibrio penaecida. An experiment of genetic selection based on the criterion ofsurvival to Syndrome 93 has been conducted at the Laboratory of Aquaculture of New Caledonia. The 3rd selected generation had demonstrated very encouraging results (survival rates improved by around 20% in experimental infections with V. penaeicida). These results have not been confirmed at the 4th generation and no difference in terms of physiology and immunology appears between the selected population and the non-selected control population. The potential causes of these results are examined and proposals for protocol improvements are given. La filière crevette de Nouvelle-Calédonie reposesur la maîtrise de la reproduction contrôlée de la crevette Litopenaeus stylirostris, espèce introduite dans les années 1980. La difficulté majeure que rencontre la filière depuis une dizaine d’années est la récurrence du « Syndrome 93 », qui s’exprime sous forme d’épisodes de mortalités lors des baisses de température aux intersaisons. Ces mortalités sont associées à la bactérie pathogène Vibrio penaeicida Une expérience de sélection sur un critère de survie à des épisodes de Syndrome 93 a été menée au Laboratoire Aquacole de Calédonie. La 3èmegénération sélectionnée avait montré des résultats très encourageants (survies améliorées de l’ordre de 20% lors d’infections expérimentales à V. penaeicida). Ces résultats ne sont pas confirmés en 4ème génération et aucune différence en termes physio-et immunologique n’apparaît entre la population sélectionnée et la population témoin non sélectionnée. Les causes potentielles de ces résultats sont examinées et des propositions d’amélioration de protocoles sont avancées. Droits : 2004 Université de la Rochelle, Ifremer http://archimer.ifremer.fr/doc/00204/31527/29943.pdf http://archimer.ifremer.fr/doc/00204/31527/ | Partager |
Proteinaceous exotoxins of shrimp-pathogenic isolates of Vibrio penaeicida and Vibrio nigripulchritudo Auteur(s) : Aguirre-guzman, Gabriel Labreuche, Yannick Ansquer, Dominique Espiau, Benoit Levy, Peva Ascencio, Felipe Saulnier, Denis Éditeur(s) : Universidad Autonoma de Baja California Résumé : The pathogenicity of two V. penaeicida strains, AM101 and KH-1, with different geographic origin, and V. nigripulchritudo strain AM102, were investigated in juvenile blue shrimp species Litopenaeus stylirostris. Alive bacteria and protein fractions (PFs) obtained from cell-free supernatants (CFS) were used in experimental challenges. Strains AM102, AM101, and KH-1 produced respectively 60, 54 and 12% mortality at 96h after infection using 104 cfu mL-1 of bacterial suspension. Exocellular toxin-like factors were evidenced in CFS from the New Caledonian strains (AM102 and AM101) but not the Japanese strain (KH-1). At 48h post injection of each CFS, mortality rates were respectively 96,98 and 5% when these strains were cultivated at 20°C, whereas only O,16 and 5% mortality rates were observed when these strains were cultivated at 30°C. Clear differences in pathogenicity between both V. penaeicida strains of distinct geographic origin (AM101 and KH-1) were thus emphasized. Protein fractions were obtained from CFS of all the strains tested in this study and cultivated at 20°C, by ammonium sulphate precipitation. Whatever the used strain, significantly higher mortalities were produced with PFs obtained with 0-40% of ammonium sulphate saturation, in respect to those produced with PFs60 and PFs80. Shrimp injected with PFs40 from strains AM102, AM101 and KH-1, and at a 20-µg org-1 dose produced respectively 100, 90 and 60% mortality 46h after the challenge. The strain AM101 showed a mediam lethal dose of approximately 5µg protein org-1 (1-1.25 µg protein g-1 body weight) 59h after injection. The PFs40 from New Caledonian strains were fond sensitive to heating and proteinase K treatments, reinforcing thus the hypothesis of their proteinaceous nature. Surprisingly, PFs40 from each bacterial strain displayed similar protein bands by SDS-PAGE suggesting that the tested strains share a common exotoxic compound regardless their distinct geographic origin or species. Ciencias Marinas (Universidad Autonoma de Baja California), 2003 , Vol. 29 , N. 1 , P. 77-88 Droits : Universidad Autonoma de Baja California http://archimer.ifremer.fr/doc/2003/publication-2807.pdf http://archimer.ifremer.fr/doc/00000/2807/ | Partager Voir aussi Pathogenicity Exotoxin Shrimp Litopenaeus stylirostris Vibrio nigripulchritudo Vibrio penaeicida Télécharger |
Quantification of Vibrio penaeicida, the etiological agent of Syndrome 93 in New Caledonian shrimp, by real-time PCR using SYBR Green I chemistry Auteur(s) : Goarant, Cyrille Merien, F Éditeur(s) : Elsevier Résumé : Shrimp farming is a small but growing industry in New Caledonia. Since 1993, "Syndrome 93" has been affecting New Caledonian shrimp farming industry every cold season, causing severe epizootic mortalities in grow-out ponds and significant losses. Highly pathogenic strains of Vibrio penaeicida are considered the etiological agent of the disease in Litopenaeus stylirostris. On one hand, studies demonstrated that healthy shrimp may carry V penaeicida for weeks with a high overall prevalence, regardless of any seasonal pattern or temperature conditions. On the other hand, larvae are free of V penaeicida and are also resistant to experimental infection. V penaeicida is frequently detected in incoming water pumped from the bays, which was shown, by a molecular typing study, to be the infectious source. This particular epidemiological pattern highlights the major role of the factors that trigger and aggravate the disease in grow-out ponds, where shrimp populations carry the pathogen all year round. In order to gain a better understanding of "Syndrome 93" epidemiology, quantification of V penaeicida both in shrimp and the shrimp farm ecosystem is necessary. This article describes the steps in the successful development of a real-time PCR quantification assay of V penaeicida in shrimp haemolymph, seawater (from ponds or bays) and sediment pore water, including the choice of an accurate extraction technique. The entire detection method; including sample processing, DNA extraction and real-time PCR amplification, can be completed within 4 h. (c) 2006 Elsevier B.V All rights reserved. Journal of Microbiological Methods (0167-7012) (Elsevier), 2006-10 , Vol. 67 , N. 1 , P. 27-35 Droits : 2006 Elsevier http://archimer.ifremer.fr/doc/2006/publication-1903.pdf DOI:10.1016/j.mimet.2006.02.013 http://archimer.ifremer.fr/doc/00000/1903/ | Partager |
DEDUCTION : A research project for shrimp farming sustainability in New-Caledonia Auteur(s) : Beliaeff, Benoit Chim, Liet Della Patrona, Luc Goyard, Emmanuel Herlin, Jose Labreuche, Yannick Walling, Emilie Ansquer, Dominique Éditeur(s) : The 11th Pacific Science Inter-Congress : Pacific Countries and their Ocean: Facing Local and Global Changes. March 2 - 6, 2009 Tahiti, French Polynesia Résumé : New Caledonian shrimp farming feasibility studies started in the mid-seventies. Today ca. 2000 tons of Litopenaeus stylirostris, introduced in 1978, are semi-intensively produced each year. Since fifteen years, production has been impacted by two seasonal vibriosis, the “Summer” syndrome caused by Vibrio nigripulchritudo in relation to pond bottom and water column quality, and the “Winter Syndrome” caused by Vibrio penaeicida in relation to drastic temperature drops . With the ambition of elucidating processes possibly leading to mortality outbreaks in ponds, Ifremer has been conducting a research program since 2003 under the DESANS project (2003-2006) and the present DEDUCTION project, equally funded by the South and North Provinces and by the Government of New Caledonia. This program has been designed in a multidisciplinary approach integrating the suspected multifactorial origin of shrimp mortalities. Environment quality in ponds (either sediment or water column) plays a major role in the shrimp ecophysiological status, influencing sensitivity to pathogens . DEDUCTION focuses on a better knowledge of phytoplankton composition and biogeochemical fluxes at the sediment-water interface in ponds. Impact of farm discharges is also investigated. In addition, a more fundamental work is conducted to understand the pathogenicity mechanisms of these two Vibrio species but also to prevent the occurrence of viral infections through a RNA interference approach. Markers of the ecophysiological status of broodstock, larvae, post-larvae and reared animals, such as enzymes testifying from an oxidative stress, are developed. Gain of performance (resistance to pathogens and growth) were obtained on hybrids of the New Caledonian and introduced Hawaiian strains. Finally the database Stylog is exploited in view of discriminating “good” from “bad” rearings on the basis of relevant indicators. Les études de faisabilité de l'aquaculture de crevettes ont débuté en Nouvelle-Calédonie dans les années soixante dix. Aujourd'hui environ 2000 tonnes de Litopenaeus stylirostris, introduite en 1978, sont produites chaque année de façon semi-intensive. Depuis quinze ans la production a été impactée par deux vibrioses : le syndrome d'été provoqué par Vibrio nigripulchritudo en relation avec la qualité de la tranche d'eau et des fonds de bassins et le syndrome d'hiver provoqué par Vibrio penaeicida en relation avec des chutes rapides de température. Dans le but d'élucider les processus entrainant des mortalités dans les élevages, Ifremer a conduit un programme de recherche depuis 2003 dans le cadre du projet DESANS (2003-2006) puis du présent projet DEDUCTION co-financé par les provinces Nord et Sud et le Gouvernement de la Nouvelle-Calédonie. Ce programme se construit dans une démarche multidisciplinaire intégrant les origines supposées multifactorielles des mortalités de crevettes. La qualité environnementale dans les bassins (les fonds et la colonne d'eau) joue un rôle majeur dans l'état physiologique des animaux, influençant leur sensibilité aux pathogènes. DEDUCTION se focalise sur une meilleure compréhension de la composition phytoplanctonique et des flux bio-géochimiques à l'interface eau/sédiment dans les bassins.L'impact des rejets des fermes est également étudié. En outre, un travail plus fondamental est conduit pour connaître les mécanismes de pathogénicité de ces deux espèces de Vibrio mais aussi pour prévenir les occurences d'infections virales par des approches ARN Interférence. Des marqueurs de l'état physiologique des oeufs, larves, post-larves et animaux en élevage, tels que les enzymes du stress oxydant sont utilisés.Les gains de performance (résistance aux pathogènes et croissance) sont obtenus sur des hybrides issus du croisement de la souche locale avec une souche hawaienne. Finalement la base de données STYLOG est exploitée dans le but de distinguer les bonnes des mauvaises pratiques sur la base d'indicateurs fiables. Droits : info:eu-repo/semantics/openAccess http://archimer.ifremer.fr/doc/00143/25438/23591.pdf http://archimer.ifremer.fr/doc/00143/25438/ | Partager |
Arbitrarily primed PCR to type Vibrio spp. Pathogenic for shrimp Auteur(s) : Goarant, Cyrille Merien, F Berthe, Franck Mermoud, I Perolat, P Éditeur(s) : Amer Soc Microbiology Résumé : A molecular typing study on Vibrio strains implicated in shrimp disease outbreaks in New Caledonia and Japan was conducted by using AP-PCR (arbitrarily primed PCR). It allowed rapid identification of isolates at the genospecies level and studies of infraspecific population structures of epidemiological interest. Clusters identified within the species Vibrio penaeicida were related to their area of origin, allowing discrimination between Japanese and New Caledonian isolates, as well as between those from two different bays in New Caledonia separated by only 50 km. Other subclusters of New Caledonian V. penaeicida isolates could be identified, but it was not possible to link those differences to accurate epidemiological features. This contribution of AP-PCR to the study of vibriosis in penaeid shrimps demonstrates its high discriminating power and the relevance of the epidemiological information provided. This approach would contribute to better knowledge of the ecology of Vibrio spp. and their implication in shrimp disease in aquaculture. Applied And Environmental Microbiology (0099-2240) (Amer Soc Microbiology), 1999-03 , Vol. 65 , N. 3 , P. 1145-1151 Droits : American society for microbiology http://archimer.ifremer.fr/doc/1999/publication-2734.pdf http://archimer.ifremer.fr/doc/00000/2734/ | Partager |
Amélioration génétique expérimentale de la crevette d'élevage de Nouvelle-Calédonie : Sélection d'une population de L. stylirostris résistante à la bactérie pathogène Vibrio penaeicida. Rapport final pour le Ministère de l'Outre-Mer Auteur(s) : Goyard, Emmanuel Goarant, Cyrille Bachere, Evelyne De Lorgeril, Julien Mugnier, Chantal Ansquer, Dominique Broutoi, Francis Brun, Pierre Résumé : The New-Caledonian shrimp industry is based on the controlled reproduction of the shrimp Litopenaeus stylirostris, a species which was introduced in the 80s. The major difficulty to which the industry has been faced for 10 years is the occurrence of the "syndrome 93", which corresponds to mortality phases when the temperature falls down in April-May-June. This mortality is associated to the pathogenic bacteria Vibrio penaecida and is expressed at different levels which are variable from year ta year and from pond to pond. No resistance to this pathology has been developed spontaneously. This is likely due to the protocole used to rear spawners, which does not allow to implement an efficient selective pressure at each generation
An experimental selection on the criteria of survival after picks of syndrome 93 has been conducted at the Laboratoire Aquacole de Calédonie. The 3rd selected generation demonstrates survival rates improved by 20% during experimental infections with V. penaeicida in comparison with a non selected control population of same genetic origin. The comparison of the correlated responses on the level of expression of 5 genes which are potentially implicated in immunity phenomena (Peneidins, lysozyme, transglutaminase. profiline, annexine) shows that the selected population has a level of expression in lyzozyme twice higher than the control population. This result suggests that the lysozyme could be a genetic marker which could be used in a selective breeding program to he developed in relation with the private hatcheries. La filière crevette de Nouvelle-Calédonie repose sur la maîtrise de la reproduction contrôlée de la crevette Litopenaeus stylirostris, espèce introduite dans les années 80. La difficulté majeure que rencontre la filière depuis une dizaine d'années est la récurrence du « syndrome 93 », qui correspond à des épisodes de mortalités lors des baisses de température en avril-mai-juin. Ces mortalités sont associées à la bactérie pathogène Vibrio penaeicida et s'expriment à des niveaux d'intensité variable d'une année à l'autre et d'un bassin à l'autre. Aucune résistance vis-à-vis de cette pathologie ne s'est développée spontanément. Ceci est vraisemblablement lié au protocole employé pour l'élevage des géniteurs qui ne permet pas d'exercer une pression de sélection efficace à chaque génération. Une expérience de sélection sur un critère de survie à des épisodes de syndrome 93 a été menée au Laboratoire Aquacole de Calédonie. La 3ème génération sélectionnée montre des survies améliorées de l'ordre de 20% lors d'infections expérimentales à V. penaeicida par rapport à une population témoin non sélectionnée de même origine génétique. La comparaison des réponses corrélées sur le niveau d'expression de 5 gènes potentiellement impliqués dans les phénomènes de défense immunitaire (penaeidine, lysozyme, transglutaminase, profiline, annexine) montre que la population sélectionnée a un niveau d'expression en lysozyme deux fois plus élevé que la population témoin. Ce résultat suggère que le lysozyme pourrait être un marqueur génétique utilisable dans un programme de sélection à développer en relation avec les écloseries de production. Droits : 2003 Ifremer http://archimer.ifremer.fr/doc/00109/22020/19643.pdf http://archimer.ifremer.fr/doc/00109/22020/ | Partager |
Pathotyping of Vibrio Isolates by Multiplex PCR Reveals a Risk of Virulent Strain Spreading in New Caledonian Shrimp Farms Auteur(s) : Labreuche, Yannick Pallandre, Laurane Ansquer, Dominique Herlin, Jose Wapotro, Billy Le Roux, Frederique Éditeur(s) : Springer Résumé : Two recurring syndromes threaten the viability of the shrimp industry in New Caledonia, which represents the second largest export business. The "Syndrome 93" is a cold season disease due to Vibrio penaeicida affecting all shrimp farms, while the "Summer Syndrome" is a geographically restricted vibriosis caused by a virulent lineage of Vibrio nigripulchritudo. Microbiological procedures for diagnosis of these diseases are time-consuming and do not have the ability to discriminate the range of virulence potentials of V. nigripulchritudo. In this study, we developed a multiplex PCR method to simultaneously detect these two bacterial species and allow for pathotype discrimination. The detection limits of this assay, that includes an internal amplification control to eliminate any false-negative results, were determined at 10 pg purified DNA and 200 cfu/ml. After confirming the effectiveness of our method using experimentally infected animals, its accuracy was compared to standard biochemical methods during a field survey using 94 samples collected over 3 years from shrimp farms encountering mortality events. The multiplex PCR showed very high specificity for the detection of V. penaeicida and V. nigripulchritudo (inclusivity and exclusivity 100%) and allowed us to detect the spreading of highly pathogenic isolates of V. nigripulchritudo to a farm adjoining the "Summer Syndrome area." This assay represents a simple, rapid, and cost-effective diagnostic tool for implementing timely risk management decisions but also understanding the seasonal and geographical distribution of these pathogens. Microbial Ecology (0095-3628) (Springer), 2012-01 , Vol. 63 , N. 1 , P. 127-138 Droits : Springer Science+Business Media, LLC 2011 http://archimer.ifremer.fr/doc/00063/17398/15169.pdf DOI:10.1007/s00248-011-9951-3 http://archimer.ifremer.fr/doc/00063/17398/ | Partager |
Acquisition of susceptibility to Vibrio penaeicida in Penaeus stylirostris postlarvae and juveniles Auteur(s) : Goarant, Cyrille Regnier, France Brizard, Raphael Marteau, Anne-laure Éditeur(s) : Elsevier Résumé : Vibriosis is a major disease problem in shrimp aquaculture, affecting all developmental stages, from larvae in hatchery tanks to juveniles and broodstock in growout ponds. However, bacterial strains responsible for vibriosis in the successive stages are usually considered to be different, and virulence specificity has been reported both at the species and at the stage levels. The so-called « Syndrome 93 » is a seasonal juvenile vibriosis caused by Vibrio penaeicida which affects Penaeus stylirostris in growout ponds and broodstock tanks in New Caledonia. This pathology does not cause any mortality in hatchery or nursery phases. An experimental infection design using balneation of postlarvae and early juveniles in V. penaeicida suspensions was used to evaluate the developmental stage at which shrimp become sensitive to this vibriosis. We demonstrated that the acquisition of susceptibility to this pathogen is very sudden and correlated with the acquisition of the definitive rostral formula, and from this draw conclusions regarding virulence mechanisms of V. penaeicida in P. stylirostris. Aquaculture (Elsevier), 1998-12 , Vol. 169 , N. 3-4 , P. 291-296 Droits : 1998 Elsevier Science B.V. All rights reserved http://archimer.ifremer.fr/doc/1998/publication-383.pdf DOI:10.1016/S0044-8486(98)00380-9 http://archimer.ifremer.fr/doc/00000/383/ | Partager |
A relationship between antimicrobial peptide gene expression and capacity of a selected shrimp line to survive a Vibrio infection Auteur(s) : De Lorgeril, Julien Gueguen, Yannick Goarant, Cyrille Goyard, Emmanuel Mugnier, Chantal Fievet, Julie Piquemal, D Bachere, Evelyne Éditeur(s) : Elsevier Résumé : Understanding of antimicrobial defence mechanisms of penaeid shrimp should help in the design of efficient strategies for the management and disease control in aquaculture. In this study, we have specifically analysed the expression in circulating hemocytes of antimicrobial peptides (AMPs) encoding genes, such as PEN2 and PEN3, ALF, crustin, lysozyme and a putative cysteine-rich peptide. We evidenced a relationship between the level of expression of some AMPs and the successful response of the shrimp, Litopenaeus stylirostris, to circumvent a pathogenic Vibrio penaeicida infection. Additionally, significant differences in some AMP transcript amounts are evidenced between control, non-selected shrimp line and the third generation breeding of shrimp selected for their survival to natural V. penaeicida infections. On the basis of these results, it will now be of great interest to determine if these AMPs are directly involved in the resistance of shrimp to infection or if they only reflect other acquired defence mechanisms which can confer a resistance. (C) 2008 Elsevier Ltd. All rights reserved. Molecular Immunology (0161-5890) (Elsevier), 2008-07 , Vol. 45 , N. 12 , P. 3438-3445 Droits : 2008 Elsevier Ltd All rights reserved. http://archimer.ifremer.fr/doc/2008/publication-4524.pdf DOI:10.1016/j.molimm.2008.04.002 http://archimer.ifremer.fr/doc/00000/4524/ | Partager Voir aussi Real time PCR Cysteine rich peptide Crustin Anti LPS factor Lysozyme Penaeidins Immune response Vibrio penaeicida Penaeid Decapoda Télécharger |
Eutrophication in a tropical pond: Understanding the bacterioplankton and phytoplankton dynamics during a vibriosis outbreak using flow cytometric analyses Auteur(s) : Lucas, Ronan Courties, C. Herbland, Alain Goulletquer, Philippe Marteau, Anne-laure Lemonnier, Hugues Éditeur(s) : Elsevier Science Bv Résumé : In tropical shrimp ponds, the increasing of feed input, concomitantly with the stocking shrimp biomass, induces an eutrophication of the ecosystem. Although difficult to maintain, its stability is required to guarantee the success of the culture. A 110-day period of phytoplankton and bacterioplankton stock and dynamics in an earthen pond (1.2 ha area, 1 m depth) was monitored using flow cytometry to provide baseline information on community characteristics and ecosystem instability. Seven autotrophic cell types were identified over the whole sampling period. Prokaryotic cells included Synechococcus sp., a group named UNK which presented an atypical new flow cytometric signature and picoeukaryotes (PEUK). Nanophytoplankton cells were represented by 4 groups: NAN1, NAN2, NAN3 and Cryptophytes. During the first part of the survey, picophytoplankton dominated the phytoplanktonic assemblage. The mean abundance of total cells (up to 8 x 10(6) cells mL(-1)) was among the highest recorded in marine and brackish waters. Bacterial abundance and production ranged from 0.8 to 5.1 x 10(7) cells mL(-1) and from 30 to 110 mu g C L-1 h(-1). A shift from pico to nanophytoplankton abundance was observed for a few days from d 96. During this period, heterotrophic bacteria production and abundance suddenly dropped, implying a change in the functioning of the microbial loop. This shift was concomitant with a significant shrimp mortality outbreak due to Vibrio penaeicida, the etiological agent of a disease known as Syndrome 93, which affects the shrimp industry in New Caledonia. This survey suggests that flow cytometric analysis could be used for the monitoring of aquaculture systems to improve our understanding of the complex phytoplankton and bacterial dynamics of these systems and its potential influence on disease development. (c) 2010 Elsevier B.V. All rights reserved. Aquaculture (0044-8486) (Elsevier Science Bv), 2010-12 , Vol. 310 , N. 1-2 , P. 112-121 Droits : 2010 Elsevier B.V. All rights reserved. http://archimer.ifremer.fr/doc/00029/14068/11312.pdf DOI:10.1016/j.aquaculture.2010.10.022 http://archimer.ifremer.fr/doc/00029/14068/ | Partager Voir aussi Tropical shrimp aquaculture Ecosystem shift Phytoplankton Bacterioplankton Vibrio Flow cytometry Télécharger |
Amélioration génétique pour la résistance au Syndrome 93 : bilan de 5 générations de sélection expérimentale Auteur(s) : Goyard, Emmanuel Ansquer, Dominique Brun, Pierre De Decker, Sophie Dufour, Robert Goarant, Cyril Patrois, Jacques Peignon, Jean Résumé : Selective breeding of shrimp for viral resistance is established for species like L. vannamei and L. stylirostris. However, selection for bacterial infection resistance is not documented. In New Caledonia, "Syndrome 93" is a vibriosis that affects cultured shrimps during the cool season with survival from 15 to 30%.
Current practises for breeders rearing at low densities do not improve the resistance of the offsprings mainly because selection pressures for "Syndrome 93" are low. Even after the fifth generation, survival improvement is not significant. La sélection génétique de lignées de crevettes résistantes à certains virus a démontré son efficacité en particulier chez L. vannamei et L. stylirostris. En revanche, la sélection de crevettes résistantes à une bactérie du genre Vibrio est peu documentée. En Nouvelle-Calédonie, le « Syndrome 93 », vibriose à V. penaeicida, affecte toutes les fermes de crevettes L. stylirostris lors des élevages de saison fraîche. Les survies en bassins de grossissement, de l’ordre de 40 à 60% dans un élevage sain, tombent à des valeurs de 15 à 30%. L’élevage de géniteurs à faible densité, s’il maximise les performances de reproduction, ne semble pas optimiser la qualité de la descendance du fait des faibles pressions de sélection qui s’exercent vis-à-vis du syndrome 93. Les améliorations significatives du nombre d’animaux survivant aux infections et aux épisodes de mortalité en bassin restent faibles en valeur absolue à la 5ème génération de sélection expérimentale en conditions d’élevage, mais sur le long terme la stratégie testée pourrait se révéler économiquement efficace. Droits : 2005 Ifremer http://archimer.ifremer.fr/doc/00117/22848/20658.pdf http://archimer.ifremer.fr/doc/00117/22848/ | Partager |
Typage moléculaire en AP-PCR des souches de Vibrio spp. isolées des épisodes de mortalité du Syndrome 93. Perspectives de lutte et conseil de gestion zoosanitaires déduits Auteur(s) : Goarant, Cyrille Résumé : Une étude de typage moléculaire par AP-PCR de souches de Vibrio impliqués dans des épisodes de mortalité d'élevages de crevettes en Nouvelle-Calédonie (Syndrome 93) et au Japon a été réalisée fin 1996 avec le concours de l' Institut Pasteur de Nouvelle-Calédonie. Elle nous a permis une identification rapide des isolats en terme de génospecies ainsi que des discriminations d' intérêt épidémiologique entre les isolats du génospecies majoritaire. Ainsi, des sous-groupes ont été identifiés au sein de l' espéce majoritaire : V. penaeicida. Certains de ces sous-groupes correspondent à des topotypes et permettent de discriminer les isolats du Japon de ceux de Nouvelle-Calédonie, ainsi que ceux de deux baies différentes de Nouvelle Calédonie, distinctes de 50 km seulement. D'autres sous-groupes ont pu être identifiés au sein de l' un des topotypes calédoniens, sans qu'il soit possible de l'expliquer par des éléments épidémiologiques. L' analyse des géotypes et des données zootechniques permettent de qualifier cette vibriose de maladie hydrique, c'est à dire dont le pathogène est véhiculé par l'eau. Des recommandations d' ordre zoosanitaires en sont déduites, qui tendent principalement à limiter les transferts d'animaux entre les différents sites d'élevage. Cette technique d'AP-PCR est utilisée pour la première fois dans l'étude épidémiologique d' une vibriose de crevettes. Sa rapidité, sa fiabilité et les informations d'ordre épidémiologique qu' elle fournit en font une technique de choix pour ce type d'étude Droits : 1997 Ifremer http://archimer.ifremer.fr/doc/00274/38541/37061.pdf http://archimer.ifremer.fr/doc/00274/38541/ | Partager |