Contribution à l'étude de virus de mollusques marins apparentés ...

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Publication 6 Diseases of Aquatic Organisms, sous presse THERMAL EFFECTS ON HERPES-LiKE VI RUS DETECTION AMONG HATCHERY-REARED LARVAL PACIFIC OYSTER, CRASSOSTREA G/GAS. R.M. Le Oeuff 1, T. Renault 1 and A. Gérard 2 1 Laboratoire de Génétique, Aquaculture et Pathologie, Unité de Recherche en Pathologie et Immunologie Générales, Abstract IFREMER, B.P.133, 17390 La Tremblade, France 2 Laboratoire de Génétique, Aquaculture et Pathologie, Unité de Recherche en Génétique et Ecloserie, IFREMER, B.P.133, 17390 La Tremblade, France This paper demonstrates thermal effects on herpesvirus·like detection in Pacifie oyster, Crassostrea gigas, larvae he Id at different temperatures. Intranuclear, intracytoplasmic and extracellular virus particles were observed in velum and mantle connective tissues of oyster larvae reared at 25-26°C. While in larvae held at 22-23°C, although showing nuclear lesions, the presence of viral particles was not detected. Results were obtained for oyster larvae with four different parental ongins. Herpesvirus infection was found in three of the four groups of oyster larvae held, in the same conditions, at the higher temperature category. Key words Herpesvirus, Pacifie oyster, Crassastrea gigas, larval culture, mortality, temperature. 100
Introduction The Pacific oyster, 'Crassostrea gigas, is the most important species of molluscs reared in the world (FAO, 1989), and is of particular importance for French mariculture. However, cultivation may be endangered by the occurrence of epizootics, especially viral diseases, which are considered one of the major risks to production. Indeed, oyster mortalities were described and associated with the presence of several families of morphologically described viruses among different species of ostreids. The first was reported in adult American oysters, Crassostrea virginica, and described as a member of the family Herpesviridae (Farley, 1972). More remarkably, mass mortalities of adult Portuguese oysters, Crassostrea angulata, among French live-stocks, were caused by an lridovirus between 1967 and 1970 (Comps and Duthoit, 1976; Comps et al., 1976; Comps and Bonami, 1977). Other important viruses, described trom ostreids, are members of the families lridoviridae, Papovaviridae, Togaviridae, Retroviridae and Reoviridae (Elston, 1979; Eiston and Wilkinson, 1985; Farley, 1976; Farley, 1978; Meyers, 1979). More recently, in 1991, other viruses belonging to the Herpesviridae, were associated with high mortalities of hatchery-reared larvae of C. gigas in France (Nicolas et al., 1992) and in New Zealand (Hine et al., 1992). Subsequently, sporadic high mortalities of larval C. gigas have been observed in 1992, 1993 and 1994 associated with a herpes-like virus, in some French hatcheries (Renault et al., 1994b). The pathogenicity of this virus was demonstrated earlier by experimental transmission of the disease to axenic C. gigas larvae (Le Deuff et al., 1994). Additionally, mortalities of oyster spat of O. edulis and C. gigas were also associated with herpes-like viruses (Comps and Co'chennec, 1993; Renault et al., 1994a). These herpes-associated mortalities among ostreids ail occurred du ring the hotter summer periods. Thus, it appeared necessary to investigate the effects of tempe rature on the induction of these viral infections. For this purpose, we cultured C. gigas larvae at high (25-26°C) or low (22-23°C) temperatures and we examined the susceptibility to the disease of larvae Irom broodstocks 01 different geographical origins. 101
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1 1 1 1 Année 1995 UNIVERSITE DE B
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à mes parents à Stéphane
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1 1 ) TABLE DES MATIERES INTRODUCTI
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3 - Essais d'infection expérimenta
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INTRODUCTION
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d'approche pour contrôler les mala
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1 1 1 1 1 1 1 1 j Un premier chapit
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DONNEES BIBLIOGRAPHIQUES 1 - Rappel
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1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1
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et Wray, 1952 ; Mackin, 1962 ; Burr
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1 1 1 1 1 1 1 1 des zones indemnes
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3 - Procaryotes Les mollusques biva
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Tableau 4 : Infections à Chlamydie
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cheptels de C. angulala, a conduit
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t t 1 1 1 1 t 1 1 t 1 1 t 1 1 t 1 1
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i 1 1 1 1 1 1 1 1 1 1 1 1 1 J 1 int
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III - Caractéristiques principales
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1 1 1 1 1 1 1 1 1 1 1 J 3.2 - Entr
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1 1 1 1 1 1 1 1 1 1 1 PREAMBULE Pub
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Chapitre 1 : Caractérisation d'un
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l 1 1 1 1 1 1 1 1 1 1 1 1 PUBLICATI
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References AllIle . w .. SchlOl fc
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Platichtys flesus, plie, Pleuronect
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Figure 1 : Marquage de coupes de do
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CHAPITRE 2 Comparaison antigénique
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Page 68 and 69: Tableau 2 : Comparaison anti géniq
Page 70 and 71: Figure 4 : Coupe histologique d'hû
Page 72 and 73: Chapitre 1 : Mise en évidence et d
Page 74 and 75: Les virions observés chez les larv
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Page 81 and 82: 2 - Participation à la mise en év
Page 83 and 84: Figure 10 : Lésions histologiques
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Page 89: 736 Material and methods Samples of
Page 93 and 94: 740 RENAULT (T.) ET COLLABORATEURS
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Page 97: Chapitre Il : Essais de reproductio
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Page 107 and 108: CHAPITRE 3 Effet de la température
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Page 121 and 122: Our plans for testing this la st hy
Page 123: LEGEND TO FIGURES Figure 1 : Origin
Page 129 and 130: CHAPITRE IV : Essais de propagation
Page 132 and 133: Figure 1 : Principales caractérist
Page 135 and 136: 2 - Essais d'infection de primocult
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Page 145 and 146: 2.2. Essais d'infection de primocul
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DJBE16 DJBE2L DJBECI DJBE16 DJBE2L
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1.2.2 - Alignement des séquences D
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viral, des spots d'intensité plus
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visualisées sur les dépôts d'ADN
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Figure 6 : Diagnostic de l'infectio
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CONCLUSION L'ensemble de ce travail
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séquences disponibles dans les ban
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REFERENCES BIBLIOGRAPHIQUES Ahne W.
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Cahour A., 1979. Mar/eilia refringe
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Darlington R.W. et Moss L.H., 1969.
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Girard M. et Hirth L., 1989b. Méth
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Johnson D.C. et Spear P.G., 1982. M
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Lemaster S. et Roizman B., 1980. He
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Ohba M., Kanda K. et Aizawa K., 199
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Read G.S. et Frenkel N., 1978. Herp
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Shuster C.L. et Hillman T.E., 1963.
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1994 : P. Maffart, R.M. Le Deuff, T
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days, according to Reed and Muench'
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detect a single infected cell in a
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Vet Res (1995) 26. 539·543 © Else
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Annexe 2 : Méthodes et techniques
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minutes, puis les coupes sont réhy
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2.1 - Préparation des solutions -
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2.6 - Coloration négative Lors des
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Gel de concentration 4% H 20 Tampon
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le tampon de transfert et superpos
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NaCI NaN) Ajuster le pH à 7 Tampon
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Epuisement des anticorps 1 - Utilis
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6 - Prélever la phase supérieure
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Tris-HCl pH8 1 ml Préparer extempo
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2 - Prélever 1 ml de tampon d'hybr
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