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We investigated the incidence of plasmid-mediated and chromosome-mediated iron uptake systems in strains of Vibrio anguillarum that belong to serotypes 01 and 02 and were isolated from different fish species and in different geographic areas. All of the strains gave positive reactions in CAS agar medium and in the Arnow test, which indicated that catechol types of siderophores were produced. The majority of V. anguillarum serotype 01 strains harbored a 65-kb plasmid similar to plasmid pJMl from strain 775, which encodes the siderophore anguibactin and its outer membrane receptor, protein OM2. All of the isolates harboring this plasmid promoted the growth of an anguibactin-deficient receptor-proficient mutant derived from strain 775, but none of these isolates promoted the growth of mutants lacking receptor OM2. Furthermore, under iron-limiting conditions all of these strains induced outer membrane proteins that were identical in size to protein OM2 of strain 775. In contrast, none of the serotype 02 strains contained a high-molecular-weight plasmid, but all of them induced the growth of mutants defective in the anguibactin-mediated system regardless of the presence or absence of receptor OM2. The serotype 02 strains, but not the plasmid-bearing serotype 01 strains, also induced the growth of Salmonella typhimurium enb-1 which utilizes only enterobactin as a siderophore. Moreover, the pattern of iron-regulated outer membrane proteins in serotype 02 isolates was totally different from the pattern in the serotype 01 strains harboring the 65-kb plasmid. Interestingly, several plasmidless strains belonging to serotype 01 behaved in the same way as serotype 02 strains in cross-feeding experiments and produced the same iron-regulated outer membrane protein patterns. These results indicate that regardless of their origins all V. anguillarum strains belonging to serotype 02 contain similar chromosome-mediated iron-sequestering systems. However, in the serotype 01 strains both chromosome-and plasmid-encoded iron uptake systems occur, but there is a significant predominance of the plasmid-mediated system.

Perkinsosis in clams in Galicia (NW Spain) is caused by the protozoan parasite Perkinsus olseni Lester & Davis, 1981. We used 5 clam species of commercial interest cultured in Galicia (Ruditapes decussatus, R. philippinarum, Venerupis pullastra, V. rhomboides, and Donax trunculus) to compare various P. olseni diagnostic techniques. Results of a nested PCR assay for the diagnosis of P. olseni were compared to those obtained using 2 classical methods of diagnosis proposed by the World Organisation for Animal Health (OIE), viz. histology and incubation in Ray's fluid thioglycollate medium (RFTM). Moreover, the same samples were analyzed by 2 separate research groups. The results obtained by PCR showed high sensitivity and good correlation between research groups. In addition, this method is faster than histopathology and incubation on RFTM and less expensive than histopathology. Moreover, nested PCR requires less specialized training for technicians than histology. Histopathology also showed high specificity and a good correlation between research groups. Results from incubation on RFTM suggest that this method could give divergent results between research groups, particularly in the case of low levels of infection, but it is nevertheless useful for disease-monitoring purposes. PCR is appropriate for rapidly screening large numbers of clams.

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Rickettsia-like organisms (RLOs) were found in the commercially farmed abalone Haliotis tuberculata in the northwestern region of the Atlantic Coast of Spain and are described from light and transmission electron microscopy observations. The RLOs measured ~1.6 × 0.9 µm and were found in intracytoplasmic, spherical to ellipsoidal vacuoles (up to 8 µm) in the epithelial cells of the digestive diverticulae. The morphological ultrastructure of these organisms was typically prokaryotic, including a plasmalemma and a thin Gram-negative type cell wall. Several ultrastructural changes were observed in the epithelial cells of the host containing the RLOs. The nuclei became pycnotic and several basophilic dense inclusions appeared in the cytoplasm. In addition, the host cell appeared lysed and was ruptured in advanced stages of infection. It was impossible to ascertain whether the RLOs are responsible for this disease, as a haplosporidian infection was also present. We can only conclude that the presence of RLOs simultaneously with a haplosporidian parasite may contribute to the mortality of the abalone host.KEY WORDS: Ultrastructure · Rickettsia-like organisms · Abalone · Spain Resale or republication not permitted without written consent of the publisherDis Aquat Org 71: [233][234][235][236][237] 2006 cultured and natural populations of Haliotis spp. from different geographic areas (Antonio et al. 2000, Bower 2004. Recently, the syndrome has been described as a severe pathogen-caused disease associated with Rickettsiales-like prokaryotes (Haaker et al. 1992, Gardner et al. 1995, Antonio et al. 2000, Bower 2004, Braid et al. 2005.The purpose of this work is to document and describe the ultrastructure of RLOs found in abalone Haliotis tuberculata. These prokaryotic parasites may contribute to the mortality of this commercially important species. MATERIALS AND METHODSJuvenile abalone Haliotis tuberculata (Linnaeus, 1758) (Gastropoda: Haliotidae) from Ireland were grown in barrels suspended from rafts in El Grove, Galicia, NW of Spain (42°29' 20'' N, 08°53' 56'' W). Fifteen specimens were observed by light microscopy (LM) during a study which aimed to identify haplosporidiosis. In addition to the haplosporidian parasite, some of the specimens were also found to be parasitized by RLOs, mainly in the epithelial cells of the digestive diverticulae.Small fragments of the parasitized tissues were fixed in 3% glutaraldehyde in 0.2 M sodium cacodylate buffer pH 7.2 for 10 h at 4°C, washed overnight in the same buffer at 4°C, and post-fixed in buffered 2% OsO 4 for 3 h at 4°C. After dehydration in an ascendant ethanol series followed by propylene oxide, the tissues were embedded in Epon. Semithin sections were stained with methylene blue-Azur II and observed under differential interference contrast (DIC; Nomarski) microscopy. For transmission electron microscopy (TEM), the tissues were ultrathin-sectioned with a diamond knife, double-stained with uranyl acetate and lead citrate and observed in a JEOL 100CXII TEM operat...