Differential effects of exposure to parasites and bacteria on stress response in turbot Scophthalmus maximus simultaneously stressed by low water depth.

The stress response of turbot Scophthalmus maximus was evaluated in fish maintained 8 days under different water depths, normal (NWD, 30 cm depth, total water volume 40 l) or low (LWD, 5 cm depth, total water volume 10 l), in the additional presence of infection-infestation of two pathogens of this species. This was caused by intraperitoneal injection of sublethal doses of the bacterium Aeromonas salmonicida subsp. salmonicida or the parasite Philasterides dicentrarchi (Ciliophora:Scuticociliatida). The LWD conditions were stressful for fish, causing increased levels of cortisol in plasma, decreased levels of glycogen in liver and nicotinamide adenine dinucleotide phosphate (NADP) and increased activities of G6Pase and GSase. The presence of bacteria or parasites in fish under NWD resulted in increased cortisol levels in plasma whereas in liver, changes were of minor importance including decreased levels of lactate and GSase activity. The simultaneous presence of bacteria and parasites in fish under NWD resulted a sharp increase in the levels of cortisol in plasma and decreased levels of glucose. Decreased levels of glycogen and lactate and activities of GSase and glutathione reductase (GR), as well as increased activities of glucose-6-phosphate dehydrogenase (G6PDH), 6-phosphogluconate dehydrogenase (6PGDH) and levels of nicotinamide adenine dinucleotide phosphate (NADPH) occurred in the same fish in liver. Finally, the presence of pathogens in S. maximus under stressful conditions elicited by LWD resulted in synergistic actions of both type of stressors in cortisol levels. In liver, the presence of bacteria or parasites induced a synergistic action on several variables such as decreased activities of G6Pase and GSase as well as increased levels of NADP and NADPH and increased activities of GPase, G6PDH and 6PGDH.

Encystment/excystment response and serotypic variation in the gastropod parasite Tetrahymena rostrata (Ciliophora, Tetrahymenidae).

Tetrahymena rostrata, which is characterized by a particular encystment-excystment cycle involving autogamy, has been recently found infecting the kidney of edible Helix aspersa snails under farming conditions. In this work, the effects of several factors on its encystment/excystment behaviour and the occurrence of different serotypes were investigated. The encystment/excystment response under starvation conditions was seriously affected by temperature. While a peak of encystment at 48 h followed by a progressive spontaneous excystment was observed at 18 and 25 °C, the encystment response was practically inhibited at 5 °C and clearly slowed down at 10 °C. At 30 °C, most of surviving ciliates remained encysted throughout the experiment, with spontaneous excystment being detected only after switching the temperature to 18 °C. Soil components also affected the encystment/excystment behaviour at 18 °C, with spontaneous excystment occurring in the presence of a sterile-filtered soil extract or mineral water but being strongly minimized with a non-filtered soil extract. Resting cysts formed in the latter extract exhibited a 3­4 times thicker and ultrastructurally more complex wall than that formed in mineral water and retained the excystment ability for about 4 weeks. Incomplete desiccation did not affect significantly the encystment response, while the mucus and kidney extracts from snails as well as a ciliate extract strongly stimulated a rapid excystment. Finally, two different serotypes infecting H. aspersa in heliciculture farms of Galicia (NW Spain) were identified, but no differences were observed between the encystment/excystment responses exhibited by two isolates belonging to each serotype.

Brachylaima aspersae n. sp. (Digenea: Brachylaimidae) infecting farmed snails in NW Spain: morphology, life cycle, pathology, and implications for heliciculture.

The life cycle of Brachylaima aspersae n. sp. (Trematoda: Brachylaimidae) in heliciculture farms is elucidated in light of field and experimental studies. Embryonated asymmetrical eggs (33.3 µm × 20.2 µm) are passed in the faeces of the definitive host, the domestic mouse (Mus musculus), and are ingested by its unique first intermediate host, the helicid snail Helix aspersa aspersa. After hatching, the miracidium develops into a highly branched sporocyst in the connective tissues of the digestive gland. Microcaudate cercariae emerging from this gastropod migrate up the ureter of the second intermediate host, the snails H. a. aspersa and H. a. maxima, and develop into non-encysted metacercariae in the kidney. Following predation of infected snails, the metacercariae develop into adults preferentially in the proximal portion of the duodenum of the definitive host. The strict oioxenic character for the first intermediate host, as well as the cercarial chaetotaxy (3 C(I)V+1 C(I)D, 10 C(II), 5 C(III)V, 14 C(III)L, 2 C(III)D, 16 H, 6 S(I), 6 S(II), 6 S(III), 2 A(I)L+1 A(I)V, 1 A(II)L, 3 ML, 1 P(I)L and 3 P(III)L), the distinct pars prostatica, the variable appearance of testes (rounded to irregular, with smooth or slightly to moderately lobulated margins), the size of eggs, the position of acetabulum (located somewhat posterior to the anterior third of body), and the microhabitat of the adult in the final host allow differentiation of B. aspersae from other well-known species in the genus. Massive infections with sporocysts or metacercariae of this brachylaimid may induce extensive pathological changes in the organs affected. Our results confirm that control of rodents in heliciculture farms is essential to minimize the potential health risks and morbimortality associated with this newly described species.

Coccomyxa sp. (chlorophyta: chlorococcales), a new pathogen in mussels (Mytilus galloprovincialis) of Vigo estuary (Galicia, NW Spain).

In this work, we describe the occurrence of irregular shaped green aggregations in the mantle, gill filaments, adductor muscle, visceral mass and haemolymph of wild mussels (Mytilus galloprovincialis) collected from the Vigo estuary (Galicia, NW Spain). Microscopic examination of these masses revealed that they consist of intracellular green algae which are spherical to oval in shape, 5 microm in length and 3 microm in width, without flagella and with a smooth surface. The algal cells present a small single nucleus, a mitochondrion, 1-2 parietal chloroplasts and lack pyrenoids. Reproduction is by formation of 2-4 autospores or daughter cells. Pigment analysis reveals the presence of photopigments typical of green algae in addition to alloxanthin, diadinoxanthin and diatoxanthin. These carotenoids are noted for the first time in a parasitic chlorophyte. The signs of infection, together with the morphological observations, suggest that this parasitic algae may be Coccomyxa parasitica. However, further molecular studies are required for confirmation. This is the first report of Coccomyxa algae parasitizing the species M. galloprovincialis.

Molecular and morphological evidence for the occurrence of Anisakis sp. A (Nematoda, Anisakidae) in the Blainville's beaked whale Mesoplodon densirostris.

Twenty-three adults (only one male) and two fourth-stage larvae of Anisakis, recovered from the stomach of a Blainville's beaked whale (Mesoplodon densirostris) stranded in Galicia (NW Spain), were studied morphologically and molecularly. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and sequencing studies carried out on the ribosomal DNA spacers and on the cox2 mitochondrial gene confirm that these nematodes belong to the taxon Anisakis sp. A, which was previously detected as a larval stage in fishes from Madeiran waters. In addition, our molecular studies demonstrate that Anisakis sp. A and the taxon Anisakis sp., previously found in other Mesoplodon spp., are the same species. The adults of Anisakis sp. A are morphologically similar to A. ziphidarum but with a shorter body length and longer spicules (right spicule 2.42 mm, left spicule 2.30 mm). Since the poor condition of the male tail cuticle prevented a proper description of the caudal plates and the pattern of caudal papillae, we propose to retain the name Anisakis sp. A until new males are described correctly.

Virus-like particles in Urastoma cyprinae, a turbellarian parasite of Mytilus galloprovincialis.

Urastoma cyprinae is a turbellarian parasite infecting the gills of several marine bivalves. We observed the presence of virus-like particles (24 to 30 nm in diameter; icosahedral symmetry) arranged in paracrystalline arrays in the cytoplasm of subepidermal dorsal cells. These particles appear to be RNA viruses, possibly related to the Picornaviridae. This is the first report of viral particles in the parasite U. cyprinae.

Eimeria species from the European pond turtle, Emys orbicularis (Reptilia: Testudines), in Galicia (NW Spain), with description of two new species.

Parasitological examination of feces from 44 Emys orbicularis from Galicia (NW Spain) revealed the presence of 2 new eimerian species, Eimeria gallaeciaensis sp. n. and E. emydis sp. n., as well as E. mitraria (Laveran and Mesnil, 1902) Doflein, 1909. Oocysts of E. gallaeciaensis n. sp. were found in 20 of 44 (45.4%) turtles and are subspherical to lightly ovoid-ellipsoid, 19.3 x 16.0 (17-22 x 15-18) microm, shape index 1.2 (1.1-1.3), with a smooth, single-layered wall. Micropyle and polar granule are absent, but an oocyst residuum is present. Sporocysts are ellipsoid, 9.7 x 5.1 (9-10 x 5-6) microm, shape index 1.9 (1.7-2.0), each with a sporocyst residuum and a conical Stieda body usually bearing 1-4 short and thin projections. Oocysts of E. emydis n. sp. were found in the feces of 5 of 44 (11.4%) turtles and are ovoid, rarely pear-shaped, 22.6 x 17.0 (20-25 x 15.5-18) microm, shape index 1.3 (1.2-1.5), with a smooth, single-layered wall with a slight thinning at the pointed end. Micropyle and polar granule are absent, and an oocyst residuum is present. Sporocysts are ellipsoid, 11.4 x 6.0 (9-13 x 5-7) microm, shape index 1.9 (1.6-2.2), each with sporocyst residuum and a prominent Stieda body bearing 3-5 club-shaped projections. In addition to the new species described, this is the first report of E. mitraria parasitizing E. orbicularis.

Parasites of swordfish from the Gulf of Guinea.

Ten parasite species were identified after a study carried out on 125 swordfish from the Gulf of Guinea waters. The samples were taken between August and November 1997; 3 of the parasite species, Caligus coryphaenae. Pelichnibothrium speciossum, and Sphyriocephalus viridis were recorded for the first time on this host.

Lymph nodes and human tumors (review).

This review examines the crucial role of regional lymph nodes (RLN) in defense against tumor progression. RLN are one of the first major components of the immune system to come into contact with tumor cells or tumor-cell products and are important in the generation of tumor-directed immune responses. Involvement of RLN by tumor cells is a prognostic index of survival and a biological indicator of a more distant metastatic disease. Enlargement of lymph nodes as a consequence of the increase in the number of lymphoid cells, is a common finding in humans. These changes of cellular organization display the most decisive evidence of the existence of an immune response within a draining lymph node. The variety of cells mediating immune response to tumors are summarized briefly. The lymphocyte subpopulations involved reflect the nature of the response and may determine the outcome of the tumor-host interaction. The composition of the lymphocyte subpopulations can be recognized in tumor-draining lymph nodes by distinctive surface-membrane markers assessable by flow cytometry. In human patients with solid tumors limited quantification of the lymphocyte subpopulations within RLN has been carried out using this technique and the results indicated that an increase in B lymphocytes in tumor-reactive lymph nodes is marked in the adenocarcinomas (colon and breast) while in other tumors, such as melanoma and squamous cell carcinoma, this increase in B lymphocytes is less pronounced. The increased number of B lymphocytes in the reactive lymph nodes suggests the existence of an immune response involving interactions between T and B cells. B lymphocytes expression of CD80 appears to increase in some reactive lymph nodes to adenocarcinomas, possibly indicating the state of activation of CD80+ B cells, and their role as antigen-presenting cells. Any improvement in the antitumor activity of RLN would be important in the immunotherapy of cancer patients. The ability to generate a large number of tumor-reactive T lymphocytes is a critical requirement for adoptive immunotherapy. Tumor-draining lymph nodes (TDLN) are an excellent source of tumor-reactive T lymphocytes and the adoptive transfer of these cells is capable of mediating the regression of tumors established both in the lung and in the brain. Although cancers elicit a vigorous immune response during the early part of their growth, the immune response is soon downregulated, permitting progressive cancer growth. Furthermore, there are date suggesting the existence of immunosuppressive mechanisms within RLN in the antitumor response. However, there are no yet conclusive data concerning the characteristics of the response or its effectiveness.