Examinando por Autor "Figueras, A"

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Assessment of in vivo effects of the Prestige fuel oil spill on the Mediterranean mussel immune system.
(Springer Nature, 2007) Ordás, MC; Albaigés, J; Bayona, JM; Ordás, A; Figueras, A
A laboratory experiment was carried out to study immune function alteration of the mussel Mytilus galloprovincialis when exposed to the Prestige oil spilled in November 2002 on the northwestern Spanish coast. Mussels were maintained for 4 months in tanks with flowing seawater and with 1, 2, and 0 kg (controls) Prestige fuel oil. Polycyclic aromatic hydrocarbon concentrations, which were determined in gills and digestive glands, were higher in digestive glands. The methylphenantrene and dibenzothiophene profiles confirmed the real exposure of mussels to the fuel oil. Immune data analysis revealed that no differences between fuel-treated and control animals were found in the cellular immune parameters measured (hemocyte viability, phagocytic activity, nitric oxide production, and chemiluminescence emission). In addition, histologic observations did not reveal tissue lesions in any of the samples, probably because of the short time of fuel-oil exposure. In contrast, significant differences were found in serum protein concentration and lysozyme activity between the fuel-treated mussels and controls. However, these humoral immune parameters were dependant on numerous environmental and physiologic factors, so it was difficult to ascertain the real effect of the fuel oil on their variability. Because hemocytes are the primary line of defense of bivalve mollusks, the results obtained in the present study suggest that the mussel immune system was not significantly affected by exposure to the Prestige fuel oil
Hemocitos de mejillón. Producción de radicales de oxígeno y nitrógeno
(Consejo Superior de Investigaciones Científicas, 2007) Novoa, B; Tafalla, C; Ordás, Camino; Figueras, A
Histopathological study of abnormal mortalities of cockle (Cerastoderma edule) in Carril (Galicia, NW Spain)
(European Association of Fish Pathologists, 2005) Ordás, MC; Figueras, A
In 1999, abnormal mortalities of cockle (Cerastoderma edule) were reported in Galicia (NW Spain). We found several potentially pathogenic organisms and cellular disorders, including neoplasic cells with a strong lysosomic activity, which could be associated with the reported cockle mortalities.
Histopathology of the infection by Perkinsus sp. in three clam species (Ruditapes decussatus, R. philippinarum and R. pullastra) from Galicia (NW of Spain)
(National Shellfisheries Association, 2001) Ordás, MC; Gómez León, J; Figueras, A
In this study, we have determined the histopathology of the infection by the protozoan parasite Perkinsus atlanticus in three exploitable clam species (carpet-shell clam Ruditapes decussatus, Manila clam R. philippinarum, and the species R. pullastra) from Galicia (NW of Spain). In histological preparations from infected animals, typical mature trophozoites of Perkinsus, containing lipid droplets and vacuole but no vacuoplast, were observed. The trophozoites were usually in groups surrounded by a light halo. Pre-palintomic tomonts, as well as palintomic tomonts with a variable number of daughter cells, were also present in the host tissues. Less frequently, immature trophozoites with an undifferentiated cytoplasm were observed. The presence of P. atlanticus cells was always associated with a strong hemocytic infiltration of the surrounding tissues. Occasionally, the parasite cells were internalized by granular hemocytes. The morphology and distribution of the P. atlanticus were similar in the three clam species studied, this being the first time that R. pullastra from Galicia has been found infected by P. atlanticus. The morphology of the parasite's life cycle stages and the histopathology of the infection resemble those reported on other species of the genus Perkinsus.
Molecular characterisation of a turbot Mx cDNA
(Elsevier, 2005) Abollo, E; Ordás, MC; Dios, S; Figueras, A; Novoa, B
The present work describes the cloning and characterisation of a full-length Mx cDNA from turbot. Expression of this gene in different organs and blood leucocytes was induced by poly I:C, in order to evaluate its capacity to regulate the Mx gene both in vivo and in vitro.
Molecular cloning and expression analysis of interferon regulatory factor 1 (IRF- 1) of turbot and sea bream
(Elsevier, 2006) Ordás, MC; Abollo, E; Costa, M; Figueras, A; Novoa, B
The interferon regulatory factor (IRF) family comprises transcription factors that regulate the expression of interferon and interferon-related cytokines. Using the RACE technique, we have determined the complete cDNA sequence of turbot (Scophthalmus maximus) and sea bream (Sparus aurata) IRFs. These sequences shared characteristics with other IRFs of fish, mammals and birds, and showed high similarity with IRF-1. Indeed, they were included in the IRF-1 cluster of the phylogenetic tree constructed with IRF-1 and IRF-2 sequences of several organisms, and presented a low number of basic amino acid residues in the carboxy-terminal end of the proteins. All of these characteristics led to the identification of turbot and sea bream IRFs as IRF-1. Two IRF-1 sequences were obtained for both turbot and sea bream, and we named them turbot/sea bream IRF-1a and IRF-1b. Turbot IRF-1a differed from turbot IRF-1b in four nucleotides. The presence of both IRF types in cDNA from 45 turbot livers was determined by RFLP, suggesting the duplication of the gene. Sea bream IRF-1b presented a deletion of 121 bp in its ORF compared to sea bream IRF-1a, and since both IRF types were present in all 25 cDNAs analyzed by PCR, we hypothesized that the truncated sea bream IRF-1b was probably an alternative splicing product. Turbot and sea bream IRF-1 expression was constitutive in every analyzed organ, as reported before for other fish species. Poly I:C significantly stimulated turbot IRF-1 expression in muscle, spleen and kidney 24 h post-treatment, while viral haemorrhagic septicemia virus (VHSV) induced a differential expression of this factor in kidney 8 h after infection. These results do not agree with those previously reported for flounder and trout IRF. Other expression experiments with turbot leukocytes stimulated in vitro with poly I:C and with brain and kidney of sea bream infected with nodavirus did not bring out differential IRF expression levels in stimulated samples with respect to controls.
Perkinsosis in molluscs: a review
(ECP Sciences, 2004) Villalba, A; Reece, KS; Ordás, MC; Casas, SM; Figueras, A
The genus Perkinsus includes protistan parasites infecting marine molluscs throughout the world, some of which are associated with mass mortalities. Life cycle involves vegetative proliferation within the host, by which a cell named trophozoite undergoes successive bipartitioning. Other stages have been observed in vitro or in vivo, depending on the species: hypnospore, zoosporangium and zoospore. Molecular taxonomy supports a close affinity between dinoflagellates and Perkinsus spp. Six species of Perkinsus are currently considered valid: P. marinus, P. olseni, P. qugwadi, P. chesapeaki, P. andrewsi and P. mediterraneus. Histology and, above all, incubation of host tissues in Ray's fluid thioglycollate medium (RFTM) are classic diagnostic methods. In addition, more sensitive and quicker molecular diagnostic techniques based on either immunoassays or PCR have been developed for Perkinsus spp. Epizootiological studies have shown a marked influence of water temperature and salinity on P. marinus infection in oysters Crassostrea virginica, thus determining parasite geographical range and temporal disease dynamics (seasonality). In vitro cultures have been established for four Perkinsus spp. Immune response to infection varies depending on host and involves phagocytosis or encapsulation of the parasite cells by host haemocytes. A polypeptide is secreted by clam Tapes philippinarum haemocytes that could kill the parasite. In vitro cultured P. marinus cells secrete proteases that are likely involved in degradation of host tissues. P. marinus can suppress the toxic oxygen radicals produced by host haemocytes. In addition to host death, sublethal effects caused by Perkinsus spp. (reduction of fecundity, growth, and condition) may have significant ecological and economic implications. Various strategies have been assayed to mitigate the consequences of P. marinus epizootics on the oyster industry: modifications of management/culture procedures, selective breeding to obtain resistant oyster strains, and the use of triploid oysters and allochthonous oyster species. Some chemotherapeutants have been proved to inhibit or kill parasite cells in vitro.
Proteolytic activity of cultured Pseudoperkinsus tapetis extracellular products.
(Elsevier, 2001) Ordás, MC; Novoa, B; Faisal, M; McLaughlin, SM; Figueras, A
Several pathogenic protozoan release proteases are necessary for host invasion and initiation of infection. We have identified proteolytic activities in extracellular proteins secreted by the clam parasite Pseudoperkinsus tapetis (Mesomycetozoa) in vitro. The protein concentration of the P. tapetis extracellular products (ECP) increased only during the first week of culture. The appearance of new proteins of 10 and 157 kDa at the second week sample and of 12 kDa at the third week sample was shown by SDS-PAGE. The protease activity rapidly increased in the first 3 weeks of culture, and five clear bands of 23, 29, 60, 67 and 96 kDa with proteolytic activity were detected in the ECP on gelatin SDS-PAGE. Using inhibitors, the proteases were identified as members of the Ca2+ dependent, serine protease family. Their optimum pH was higher than pH 9.4. The protease activity of the P. tapetis ECP was different than that described for Perkinsus marinus, an oyster pathogen very similar morphologically to the clam parasite and member of the genus in which P. tapetis had been initially included.
Toxin and molecular analysis of Gymnodinium catenatum (Dinophyceae) strains from Galicia (NW Spain) and Andalucía (S Spain)
(Oxford University Press, 2004) Ordás, MC; Fraga, S; Franco, JM; Ordás, A; Figueras, A
Nineteen strains of Gymnodinium catenatum were isolated from one bloom in Andalucía (S Spain) and from different blooms in Galicia (NW Spain). The PSP toxin profiles of 16 of the strains were analyzed, and although the saxitoxin was exclusive to the Galician strains, the corresponding dendogram showed no clustering of the isolates from this location. However, nine out of eleven Andalusian strains were included in the same cluster. In order to compare toxin with molecular analysis, a fragment of the large subunit ribosomal (LSU) RNA gene was partially sequenced for all of the strains and fully sequenced for the five strains that had shown most different growth curves. Since all the strains were identical in the LSU sequenced region, another fragment comprising the internal transcribed spacer 1 (ITS 1), 5.8S rRNA gene and ITS 2, was sequenced and compared among all the strains. Although this region has been used before for the detection of intraspecific variability, it was similar in all our strains. Finally, to detect molecular differences in the strains, a random amplified polymorphic DNA (RAPD) analysis was performed. The corresponding cluster analysis grouped the strains in three clusters: one of them comprised all the Galician strains plus three from Andalucía, another one included eight Andalusian strains, and the last one, more separated from the two previous, was constituted by two Andalucía isolates. Although the results of the toxin and RAPD analysis were different, seven Andalusian strains were clustered together in both dendograms. Since neither the toxin nor the RAPD analysis brought out a clear geographic signal, we can conclude that differences in toxin content and RAPD profile between the isolates of G. catenatum are probably not linked to the location in which the strains were collected.
Turbot TNFα gene: Molecular characterization and biological activity of the recombinant protein
(Elsevier, 2007) Ordás, MC; Costa, MM; Roca, FJ; López-Castejón, G; Mulero, V; Meseguer, J; Figueras, A; Novoa, B
The tumor necrosis factor (TNF) superfamily is composed by several proteins with similar structure and functions. One of the main representatives of this family is TNF-alpha (TNF ), a proinflammatory cytokine which is produced by different immune cells and presents a wide variety of activities. Using the RACE technique, we have cloned and sequenced the turbot TNF cDNA. The analysis of its sequence showed several conserved motifs characteristic of members of the TNF family. A phylogenetic tree constructed with different TNFs of fish and mammals grouped our sequence within the fish TNF cluster. Therefore, the turbot TNF here studied was identified as TNF . The complete TNF gene was obtained by gene walking, and, similarly to the other known fish TNF genes, presented three introns and four exons. A PCR was designed to study the turbot TNF expression in vivo using as stimulus the bacteria Vibrio pelagius strain Hq222 and virus VHSV. The expression of the cytokine happened early after injection, and it was dependent on the pathogen injected and organ analyzed. Virus induced a higher TNF expression, but this response was shorter in time than that induced by bacteria. In addition, TNF expression was in general higher in kidney than in liver, as expected since the former is the haematopoietic organ of fish. The turbot recombinant TNF (rTNF ) was obtained by IPTG induction of bacteria transformed with the pET15b-TNF construct, and it was purified in native conditions. The recombinant protein was approximately 20 kDa in size, and its biological activity was assessed in vitro. No effect of the rTNF neither alone nor in combination with LPS was observed on the chemiluminescence activity of turbot macrophages at any time tested. However, NO production was enhanced by the recombinant protein alone or with LPS 72 h after the addition of the treatments. Finally, turbot rTNF was able to recruit and activate inflammatory cells when injected in gilthead seabream, although to a lesser extent than gilthead seabream rTNF .