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

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HERPESVIRUSES ASSOCIATED WITH MORTALITIES AMONG PACIFIC OYSTER, IN FRANCE Host species Size References Capsids Enveloped virions C. virginiea 70/90 nm 1 161 C. gigas 70±2.Dnm 90 ± 5.0 nm 1121 C. gigas 97 ± 4.0 nm 131 ± 9.0 nm 1101 O. edulis BOnm 160/1BOnm 141 C. gigas (lafVae) 75 ± 3.0 nm 122 ± 6.0 nm ln this report C. gigas (young spats) 75 ± 3.0 nm 121 ± 7.0"m ln thi5 report TABLE IV. - Morphological characleristics of herpcs-like viruses observed among bivalve molluscs. Indeed, it seems that the herpes-Iike viruses detected among bivalve molluscs are not a real danger for animaIs cultured at low temperatures Iike the observations performed by FARLEY el al. [10] suggest. Thus, since 1970, no catastrophic mortalities associated with herpes-like virus detection were reported among the species C. virginica. However it seems necessary to perform regular controls for a survey of the evolution of these virus infections among C. gigas oysters. The evolution of the disease in hatchery-reared larvae with the flfst signs visible three to four days after the spond seems to point out that contamination of larvae occurs quite carly, and may result of a vertical transmission. Among the batches of 3-7 month old young oysters, the mortalities appcared suddenly and the course of the disease was very brief in time, only a week. These reports suggest a high pathogenicity of this herpes-Iike virus, with a great infectivity in a same batch. But in the case of young spat oysters, the origin of virus contamination is uncJear. Moreover, the presence of two similar viruses among C. gigas larvae and C. gigas young oyster spat may indicate that some larvae, survivors of animais infected by herpes Iike viruses could be healthy latent carriers and express subsequently the disease under stressed conditions. In point of fact, four batches of herpes-Iike virus infected C. gigas young oyster spat are originating from French hatcheries and the larvae were produced in 1993. One bateh of these infected young oysters came from a hatchery in which mortalities in association witl! herpes-virus detection were observed among several breedings during the summer of 1993. The three other batehes were originating from hateheries in which no larvae sam pie was taken and no examination was made during their productions, So it is impossible to know if the larvae were infected or not by the herpes-Iike virus before their exit of these hateheries. Purification· and fine characterization seem to be necessary to develop sensitive diagnosis methods for detection of these pathogens described among C. gigas larvae and C. gigas young oysters and to determine relationship between these viruses. 84 Acknowledgements This sludy could nOI been completed without the valuable help of sorne oyster French farmers and priva te French hatcheries. The aUlhors ack. nowledge André OIË.RARD for providing biological material. Thanks to D' Henri GRIZEL for critical comments on the manuscript. This research \Vas supported in part by the Conseil Général de la Charente-Maritime. References 1. - AWERMAN (D.J.) : Shellfish diseases, past, present and future. Ill: Proc. I1th Ann. Conf. Sellfish Assoc. Great Brilain, 1980, p. 30-40. 2. - BUCHANAN (J.A.) and RICHARDS (R.H.) : Herpes-like virus diseases of marine organisms. Proe. R. Soc. Edimb., 1982,810, 151·168. 3. - CAMPADELU-FuIME (O.), FARABEGALI (F.), DI GAETA (S.) and ROIZMAN(B.) : Origine of unenveloped capsids in the cytoplasm of eells infected with Herpes Simplex Virus I. J. Virol., 1991, 65, 1589-1595. 4. - COMPS (M.) and DUTHolT (J.P.) : Infection virale associée à la « maladie des branchies» de l'huître portugaise, Crassostreo angula/a LmK. CR. Acad. Sei., 1976,283 D. 1595-1596. 5. - COMPS (M.), BONAMI (J.R.), VAGO (C.) and CAMPILLO (A.) : Une virose de l'huître portugaise (Crassos/rea angulala LMK). C.R. Acad. Sei., 1976, 282 D, 1991·1993. 6. - COMPS (M.) and BONAMI (J .R.) : Infection virale associée à des mortalités chez l'Huître Crassostrea gigas Thunberg. C.R. Acad. Sei., 1977,285 D, 1139-1140. 7. - COMPS (M.) and COCHENHEC (N.) : A herpes-like virus from the European oyster Os/rea edulis. J. [nver/ebr. Pothol., 1993,62, 201-203. 8. - DARGAN (D.l.) : The structure and assembly of herpes-viruses. 1" : Electron microscopy of proteins, 5 Viral structure. J .R. HARR IS (ed). Academic Press, New York, 1986, p. 359-437. 9. - ElSTON (R.) : Virus-Iîke partides associated with lesions in larval Pacific oysters (Crassos/rea gigas). J./nvertebr. Palho!.. 1979. 33,71-74. 10. - FARlEY (C.A.), BANFIElD (W.O.), KAsNIC (G.) and FOSTER (W.S.) : Oyster Herpes-type virus. Science. 1972, 178,759-760. Il. - FURLONG (O.), SWIFT (A.) and ROIZMAN (B.) : Arrangement of herpes virus deoxyribonucleic acid in the core. J. V;rol., 1972, 10, 1071-1074. 12. - HEDRICK (R.P.), GRAFF (J.M.), OKJHIRO (M.S.) and Mc DOWELL (T.S.) : Herpes viruses detected in papillomatous skin growths of Koï carps (Cyprinus carpio). J. Wild. Dis., 1990, 26, 576-581. 13. - HEDRICK (R.P.) and SANO (f.) : Herpesviruses of fish. ln : Viruses of lower vertebrales. V. AHNE, E. KURSTAK (eds). SprÎnger Verlag, Berlin, 1989, p. 161 -170. 14. - HINE (P.M.), WESNEY (B.) and HAY (8.E.) : Herpesvirus associated \Vith mortalities among hatchery-reared larval Pacific o)'ster Crassosfrea gigas. Dis. aqllat. Org., 1992, 12, 135-142. 741
742 15. - MURP HY (F.A.) and K INGS BURG (D.W,) : ln: Virus taxonomy. Vi rology. B.N . FIELDS. D.M . KNIPE (eds). Raven Press, New York, 1990, p. 9-35. 16. NICOLAS (J.L.), CoMPS (M.) and COCHENNEC (N.) : Herpes-like virus infec ting Pacifie oyster larvae, CrassaS/feo gigas. Bull. Eur. Ass. Fis" Pot/loI., !992, 12, 11·13. 17. ROIZMAN (B.) : /11 : Herpesviridae : a brier introduction. Viro (ogy. B.N. FIELDS, D.M. KNIPE (eds.). Raven Press, New York, 1990, p. 1787-1 793. 18. RorzMAN (B.), CARMICHAEL (L.E.), DEIN HARDT (F.), D E THE (G.), NAHMIAS (A.), PLOROUGHT (W.), RAPP (F.), SHELDR ICK (P.), TAKAHASHI (M.) and WOLF (K.) : Herpesviridoe. I ntervÎ· rology, 1981, 16, 201 -2 17. 85 RENAULT (T.) ET COLLABORATEURS 19 . - SAND (T.), FUKUDA (H.) and FURUKAVA (M.) : Herpesvirus cyprin;: biological and oncogenic properties. J. Fis" Path., 1985 , 20, 38 1-388. 20. - SEVERI (B.), LANDIN I (M.P.) and GOVONI (E.) : Human cylomegabvirus morphogenesis an ultrastructural study of the laIe cytoplasmic phases. Arch. Viro/., 1988,98,5 1-64. 21. - TRALKA (T.S.), CoSTA (J.) and RALSON (A.) : Electron microscopie study o f herpes-virus saimiri. Vir% gy, 1(jJ7, 80, 158- /65. 22. - TuMILOWICZ (J.J.) and POWELL (B. B.) : Raie o f intranuclear sacs, intrasaccatc tubes, and cyloplasmic dense bodies in the structura i complelion of cytomegalovirus. IlIlervirology, 1990, 3 t. 14-22.
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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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1 1 1 1 1 1 1 ! 1 1 1 1 ! 1 1 1 1 1
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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 1 1 1 1 1 1 1
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Page 45 and 46: 1 1 1 1 1 1 1 1 1 1 1 PREAMBULE Pub
Page 47 and 48: Chapitre 1 : Caractérisation d'un
Page 49 and 50: l 1 1 1 1 1 1 1 1 1 1 1 1 PUBLICATI
Page 52 and 53: References AllIle . w .. SchlOl fc
Page 55 and 56: Platichtys flesus, plie, Pleuronect
Page 57: Figure 1 : Marquage de coupes de do
Page 60 and 61: CHAPITRE 2 Comparaison antigénique
Page 62 and 63: Ces électrophorèses ont égalemen
Page 64 and 65: 1\6 , HMW LMW Figure 1 : Caractéri
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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
Page 85: Bu ll. Eur. Ass. Fi sh Palhal.. 14(
Page 89: 736 Material and methods Samples of
Page 93: 740 RENAULT (T.) ET COLLABORATEURS
Page 97: Chapitre Il : Essais de reproductio
Page 103 and 104: 2 - Essais d'infection expérimenta
Page 107 and 108: CHAPITRE 3 Effet de la température
Page 109 and 110: 26°C), de détecter la présence d
Page 111 and 112: présentent des lésions histologiq
Page 113: Introduction The Pacific oyster, 'C
Page 117 and 118: (Table 1). 100% mortality in this g
Page 119 and 120: than at high temperature (25-26°C)
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
Page 137 and 138: (SVF) et/ou d'hémolymphe de Crassa
Page 139: 68 0.2 g/I, KCI, No. P 1300' 2.77 g
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2.2. Essais d'infection de primocul
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Chapitre V : Mise au point d'un pro
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Du fait de la présence d'une coqui
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• " (1 - " 100nm 100nm Figures 1
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Chapitre VI : Mise au point de mét
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chromatine est rejetée en périph
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528 RM Le DeuN et al ,GD Fig 2. Imm
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1.2 - Recherche de séquences conse
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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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