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1.
Three species of myxobolid parasites were found infecting the gills of the Iberian nase, Chondrostoma polylepis , in the River Ulla (Galicia, NW Spain), Myxobolus leuciscini , M. impressus and M. gallaicus sp. nov., which is described herein. Myxobolus leuciscini locates in the intralamellar space and the plasmodium begins to develop in the endothelium of the capillaries of the secondary lamellae. Myxobolus impressus plasmodia are located in the interlamellar multilayered epithelium. Myxobolus gallaicus plasmodia are elongated and are located exclusively in the central venous sinus of the branchial filament, suggesting that they begin their development in branchial endothelial tissue. We also present data on spore size and morphology, plasmodium and pansporoblast characteristics and pathological effects of the three species in the host branchial tissues.  相似文献   
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Research on intraspecific variation in ciliates is scarce, and in scuticociliate parasite of fish, virtually nonexistent. In this study, seven isolates obtained from turbots affected by scuticociliatosis in different parts of the Iberian Peninsula (northwest Spain and southwest Portugal) were morphologically and genetically characterized to investigate the intraspecific divergence in these amphizoic ciliates. The isolates were stained with ammoniacal silver carbonate and examined in an optical microscope; all were found to have the typical morphological characteristics described for Philasterides dicentrarchi (syn. Miamiensis avidus). Sixteen biometric characteristics of the seven isolates were used in a canonical discrimination analysis (CDA) to select a subset of those that best identified each isolate. Discriminant analysis indicated that the OPK3 width, length of the PM2, length of the buccal field, the body width, L:W ratio, the body length, the OPK1 width and the distance between OPK2 and OPK3 were the most important morphological variables for discriminating the isolates. The first three canonical functions accounted for 86% of the total variance. The scatter plots of the first two canonical variables grouped and separated the P. dicentrarchi isolates into five clusters. Flow cytometry analysis of isolates also indicated intraspecific polymorphisms among P. dicentrarchi isolates. Nuclear markers (a 349-bp and a 390-bp fragment of 18S rRNA and β-tubulin genes) and a 398-bp of the mitochondrial cytocrome oxidase subunit I (Cox1) gene were then used to investigate the intraspecific genetic variation in P. dicentrarchi. Haplotype analysis and neighbour-joining phylogenies of nucleotide sequences of seven isolates revealed a high degree of intraspecific genetic variation among the isolates. Analysis of Cox1 and β-tubulin genes revealed six haplotypes (and clusters) in both cases; however, analysis of the 18S rRNA gene revealed only two haplotypes. The results show clear intraspecific variation at morphological and genetic levels in the scuticociliate P. dicentrarchi, and verify the suitability of mitochondrial (Cox1) and nuclear (β-tubulin) genes for detecting intraspecific genetic variation within populations of scuticociliates that infect cultured turbot. The existence of this intraspecific variation must be taken into account in the design of an effective vaccine to control scuticociliatosis.  相似文献   
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Philasterides dicentrarchi is a scuticociliate causing fatal disease in farmed turbot and sea bass. In view of its high virulence and endoparasitic location, this parasite cannot be effectively controlled by formalin baths, and no systemic chemotherapeutic treatments have yet proved effective; immunoprophylaxis may thus be an attractive alternative approach. Since vaccine development is greatly facilitated by axenic culture of the pathogen, we have developed a simple axenic culture system based on commercially available Leibovitz L-15 medium, supplemented with fetal bovine serum, lipids (lecithin and Tween 80), nucleosides and glucose. After 1 week's culture under optimal conditions (salinity 10 per thousand, pH 7.2, temperature between 18 and 23 degrees C), yields of 1-2 x 10(5)cells/ml were obtained. Even cultures with seeding densities as low as 20 cells/ml were found to produce a good yield of ciliates (about 6 x 10(4)cells/ml) after 11 days of incubation. The ciliates thus obtained were free of contamination by other microorganisms, enabling preparation of pure P. dicentrarchi antigens for vaccine development studies.  相似文献   
6.
The scuticociliatosis produced by the endoparasite Philasterides dicentarchi is a severe parasitic infection of farmed turbot (Scophthalmus maximus) characterized by several histopathological effects including extensive inflammation. Indomethacin is a nonsteroidal anti-inflammatory drug that specifically inhibits synthesis of the proinflammatory mediator prostaglandins. The effect of indomethacin on the in vitro growth of P. dicentrarchi was investigated. In vitro growth of the scuticociliate was significantly inhibited by treatment with 100 microM indomethacin for 48 h. Higher concentrations of indomethacin (mM levels) did not affect the gelatinolytic activity of the cysteine proteinases of P. dicentrarchi. In vitro treatment with 25, 50 or 100 microM indomethacin for 3 days did not significantly affect the enzymatic activity of cysteine proteinases, as assayed with p-nitroanilide as substrate. Immunoblot analysis with anti-cysteine proteinase antibodies revealed an increase in proteinase expression (molecular weights of 80, 32 and 40-45 kDa) in parasite lysates originating from in vitro cultures incubated with 25 microM indomethacin for 72 h. Degradation of genomic DNA of the ciliates was observed in cultures incubated with 100 microM indomethacin for 1, 3 and 7 days. The results suggest that indomethacin is capable of inhibiting in vitro growth of the scuticociliate P. dicentrarchi by a mechanism related to the induction of programmed cell death, without affecting the enzymatic activation of parasite proteinases, which demonstrates the potential therapeutic use of this drug in the control of turbot scuticociliatosis.  相似文献   
7.
The histiophagous scuticociliate Philasterides dicentrarchi is the aetiological agent of scuticociliatosis, a parasitic disease of farmed turbot. Curcumin, a polyphenol from Curcuma longa (turmeric), is known to have antioxidant and anti‐inflammatory properties. We investigated the in vitro effects of curcumin on the growth of P. dicentrarchi and on the production of pro‐inflammatory cytokines in turbot leucocytes activated by parasite cysteine proteases. At 100 μm , curcumin had a cytotoxic effect and completely inhibited the growth of the parasite. At 50 μm , curcumin inhibited the protease activity of the parasite and expression of genes encoding two virulence‐associated proteases: leishmanolysin‐like peptidase and cathepsin L‐like. At concentrations between 25 and 50 μm , curcumin inhibited the expression of S‐adenosyl‐L‐homocysteine hydrolase, an enzyme involved in the biosynthesis of the amino acids methionine and cysteine. At 100 μm , curcumin inhibited the expression of the cytokines tumour necrosis factor‐alpha (TNF‐α) and interleukin‐1 beta (IL‐1β) produced in turbot leucocytes activated by parasite proteases. Results show that curcumin has a dual effect on scuticociliatosis: an antiparasitic effect on the catabolism and anabolism of ciliate proteins, and an anti‐inflammatory effect that inhibits the production of proinflammatory cytokines in the host. The present findings suggest the potential usefulness of this polyphenol in treating scuticociliatosis.  相似文献   
8.
An amoeba associated with a severe gill disease affecting farmed turbot, characterized by pronounced hyperplasia of the branchial epithelia and accumulation of mucus between the secondary lamellae, is described for the first time. The amoeba was isolated, cloned and cultured on Bactoagar. Light microscopy showed that trophozoites had a mean length of 12 μm (range 8–20 μm), with a nucleus 3–5 μm in diameter containing a large central nucleolus 1–3 μm in diameter. Transmission electron microscopy showed that the mitochondria, Golgi bodies, rough endoplasmic reticulum, microfilaments, cytoplasmic microtubules and glycocalyx were similar to those previously reported for the genus Platyamoeba. These and other findings strongly suggest that the amoeba isolated is of the genus Platyamoeba, while its morphometric characteristics suggest that it may represent a new species. This is the first report of a species of the genus Platyamoeba in association with gill disease in turbot.  相似文献   
9.
The efficacy of a vaccine against the fish pathogen Philasterides dicentrarchi was evaluated in turbot by measuring the production of specific antibodies and duration of protection. Four groups of turbot were vaccinated twice, on days 0 and 30, with phosphate-buffered saline, mineral oil adjuvant, antigen or antigen plus adjuvant. Specific serum antibodies were determined on day 0 and 1 month after the first and the second vaccinations. Protection was evaluated 1 month after the first vaccination and 1 and 5 months after the second vaccination. Serum antibody titres, measured by enzyme-linked immunosorbent assay, and protection, assessed by challenges, increased significantly 1 month after the second vaccination in the group injected with antigen plus adjuvant and the protection lasted for at least a further 5 months in this group. The relative protection was 77% and 66% 1 and 5 months after the second vaccination, respectively. Administration of antigen or adjuvant separately had no effect on antibody response or protection. The results indicate that emulsion containing antigen plus adjuvant induced durable protection against P. dicentrarchi after the administration of the two vaccinations, and that this preparation can be used as a vaccine against the pathogen.  相似文献   
10.
We investigated non-specific responses of turbot spleen- and pronephros-resident adherent cells to spores of fish microsporidians, and the effects of the glucocorticoid dexamethasone (DX) on these responses. On average, 65% of adherent cells from the spleen and pronephros showed esterase activity (as characteristic of macrophages); 32% showed peroxidase activity (as characteristic of neutrophils), and 19% of peroxidase-positive cells were capable of phagocytosing microsporidian spores. A significantly higher proportion of adherent cells showed phagocytic activity when viable spores were the target than when non-viable spores were the target. Microsporidian spores stimulated adherent cells to produce reactive oxygen and nitrogen intermediates (ROIs and RNIs), though less effectively than the other stimulants tested. Adherent cells exposed to viable spores produced significantly less intracellular superoxide than adherent cells exposed to non-viable spores. Daily injection of fish with DX over 6 days significantly inhibited both phagocytosis of microsporidian spores and spore-induced ROI production, and similar effects were observed when adherent cells were exposed to DX in vitro.  相似文献   
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