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Bacterial cells of the marine fish pathogen Photobacterium damsela subsp. piscicida were grown in novel culture media. A mixture of whole cells and extracellular components was inactivated and used in bath, intraperitoneal (i.p.) and oral vaccination of sea bass, Dicentrarchus labrax, employing two sizes of fish. A commercial vaccine was used for comparative purposes. Control and immunized fish were either bath or intraperitoneally challenged 6 and 12 weeks post-vaccination. Small fish had significantly higher relative percentage survival with the novel vaccine mixture both at 6 and 12 weeks post-vaccination by bath, in comparison with the commercial vaccine. No protection was afforded at 6 or 12 weeks post-immunization by either vaccine after challenge via i.p. injection. Sea bass (1.5-2 g) intraperitoneally vaccinated with various adjuvanted vaccine mixtures were not protected against pasteurellosis. In contrast, larger sea bass (20 g) benefited from vaccination with the novel vaccine mixtures. Intraperitoneal challenge with the pathogen resulted in protection in both fish groups vaccinated with novel vaccine mixtures, whereas control fish suffered high mortalities (> 80%). Orally vaccinated fish were immersion challenged with the pathogen. At 6 and 12 weeks post-vaccination the control fish had a high mortality and the fish vaccinated with the novel vaccine mixture achieved good protection.

Photobacterium damsela ssp. piscicida (Phdp) isolates were grown in various bacteriological media, in eukaryotic cell culture media and in the presence of fish cells (resembling some aspects of in vivo growth environments). Bacterial cells, extracellular products (ECPs) and crude capsular polysaccharide were isolated and analysed by electrophoresis and Western blot using sea bass sera. Growth in bacteriological media conserved the synthesis of cell and extracellular components when these were compared with those prepared under near-in vivo growth conditions. In fact, synthesis of a larger range of cell components was induced after growth in bacteriological media. Certain media based on yeast extract and peptones from various sources and a specific salt formulation induced the synthesis of novel cell components at approximately 21.3 and 14 kDa. These antigens were recognized by sea bass sera collected after natural pasteurellosis outbreaks and other sea bass sera raised against live or inactivated Phdp cells. The ECPs of the pathogen were not good immunogens in their soluble form despite various treatments prior to immunization. The results are discussed with respect to vaccine development.

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The biochemical properties and antibiotic sensitivity patterns of 13 Pasteurella piscicida isolates from Greece are described and compared with 10 Japanese and five European (Italian and French) isolates. Morphologically and biochemically, all isolates of P. piscicida tested were nearly identical and only minor differences could be detected with reference to the level of acid production from sugars (especially from glucose). Greek, Italian and French isolates showed similar antibiotic sensitivity patterns, being resistant to erythromycin, kanamycin and streptomycin, and sensitive to most of the other antibiotics tested. Resistance was also demonstrated for some isolates towards the potentiated sulphonamides. The Japanese isolates appeared to be resistant to a larger range of antibiotics including erythromycin, kanamycin, streptomycin, oxolinic acid, oxytetracycline and chloramphenicol. Multiple resistance of Japanese strains was also noted. These results are discussed in the hght of future measures for the control of pasteurellosis.

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The effect of iron limitation, using the iron‐chelating agent 2,2 dipyridyl, on the electrophoretic profiles of outer membrane proteins (OMPs) and extracellular products (ECPs) from 21 Pasteurella piscicida strains isolated from Europe and Japan was investigated. In addition, the effect of iron‐limited and iron‐surplus growth conditions on caseinase activity in culture supernatants of the pathogen was examined. The majority of P. piscicida strains, from Greece, Italy and France, cultured under iron‐limited conditions, produced four novel OMPs (63 and three at and above 200 kDa). In contrast, iron‐regulated outer membrane proteins were not induced in Japanese strains. Electrophoretic analysis of the ECPs from the pathogen grown under iron surplus and iron limitation revealed a large range of products and additional high molecular mass (MM) bands were evident under iron‐limited conditions. When culture supernatants were analysed for their activity, most of the bacteria tested showed elevated activities under iron limited conditions. Finally, neither hydroxamate nor phenolate type siderophores could be detected with any of the chemical assays used.