Prostaglandins E2 and D2-regulators of host immunity in the model parasite Diphyllobothrium dendriticum: An immunocytochemical and biochemical study.

The spectrum of immunomodulating molecules produced by tapeworms is not yet well understood. The aims of this study, on the tapeworm Diphyllobothrium dendriticum, were: 1) detection and quantification of prostaglandins (PGs) E2 and D2 by high performance liquid chromatography; 2) visualization of PGE2 and PGD2 in specific cells, using methods of immunocytochemistry and confocal laser scanning microscopy; and 3) investigation of the ultrastructure of the cells potentially producing PGE2 and PGD2. The PGE2 immunoreaction (IR) was found in the apical terminals of the frontal glands and sensory organs in the tegument and in small neurons belonging to the main cords and commissures. PGE2-IR partly coincided with α-tubulin-IR. PGD2-IR occurred in the muscle fibers of longitudinal and transverse body muscles and coincided with phalloidin TRITC staining. Both PGE2 and PGD2 were found in the flame cells of the excretory system. Ultrastructural study of the tegument revealed two types of structures that potentially produce PGE2: ciliated and unciliated free nerve endings and frontal gland terminals reinforced with neurotubules. In the main nerve cords, small neurons were identified as potentially exhibiting PGE2-immunoreactivity. In homogenates of the plerocercoids, the measured content of PGE2 and PGD2 was 33.15ngmg-1 and 1.94ngmg-1 of fresh tissue weight, respectively. We found evidence of PGE2 and PGD2 in D. dendriticum parasitizing Coregonus autumnalis (fish) and proved excretion of PGE2 and PGD2 in response to C. autumnalis blood serum. Prostaglandins produced by D. dendriticum probably play a dual role: 1) PGE2 and PGD2 potentially modulate the fish antiparasitic immune response; 2) PGE2 is presumably necessary for proper development and function of the nervous system, and PGD2 can act as an antagonist against mediators causing muscle contraction.

[The reaction of the bursa and thymus eosinophils in the herring gull after the experimental infection with gull-tapeworm].

Using the methods of light microscopy, eosinophil topography, quantitative and qualitative changes (degranulation level of and a cationic protein content) were studied in the thymus and bursa of 36 herring gull nestlings Larus argentatus mongolicus (Suskin, 1925) 2 weeks after experimental infestation with gull-tapeworm Diphyllobothrium dendriticum (Nitsch, 1824). Eosinophils in the thymus were located in trabecules, mainly close to the blood vessels, thymic (Hassall's) corpuscles and also directly inside them, while in the bursa they were found within the internodular space. As compared with the control bird counts, relative eosinophil count in the birds with an average invasion intensity was increased 3.8 times in the thymus and 2.5 times in the bursa. In birds with high invasion intensity, these counts were increased 4 times in the thymus and 1.2 times in the bursa.

Modulation of granulocyte responses in three-spined sticklebacks Gasterosteus aculeatus infected with the tapeworm Schistocephalus solidus.

Leukocytes isolated from the head kidney and peripheral blood of 3-spined sticklebacks Gasterosteus aculeatus L. were analysed by means of flow cytometry during infection with the tapeworm Schistocephalus solidus (Müller, 1776). Although parasites increased their body weight continuously throughout the observation period (98 d), proportions of granulocytes increased in blood and head kidney only up to Day 63 post-infection (p.i.). Thereafter, declining proportions of granulocytes were observed in both organs. Thus the relative decrease in granulocyte number was not correlated to a decline in the parasitic load of the fish. To investigate a possible modulatory impact of S. solidus on granulocyte function, head kidney leukocytes were isolated at times before Day 63 p.i. and tested in vitro for their capacity to produce reactive oxygen species (ROS). Head kidney leukocytes from S. solidus-infected fish, analysed immediately after isolation (ex vivo, Day 40 p.i.), exhibited a higher ROS production when stimulated with phorbol myristate acetate (PMA), than leukocytes from naive, sham-treated control fish and fish that had resisted or cleared the infection (exposed but not infected). The latter showed an increased spontaneous ROS production that was not correlated to the numbers of granulocytes present in the head kidney isolates. In infected sticklebacks, spontaneous and PMA-induced ROS production was significantly correlated with numbers of granulocytes present in the head kidney isolates, suggesting that elevated ROS production was due to higher numbers of responding cells rather than an increased capacity of single cells. In vitro, after cultivation for 4 d in the presence of pokeweed mitogen (PWM) or extracts from S. solidus, head kidney leukocytes from control fish showed an increased ROS production and phagocytic activity compared with non-stimulated control cultures. In contrast, head kidney leukocytes from infected fish isolated on Days 48 and 44 p.i., failed to respond to S. solidus antigens in vitro. During S. solidus infection, granulocyte mobilisation resulted in elevated numbers of these cells in head kidneys, but the lack of an in vitro response to S. solidus antigens indicates an in vivo priming of granulocytes by the parasite. These observations may reflect the ability of S. solidus to impair the host's immune response once the parasite is developing in the body cavity of G. aculeatus.

Sequential development of the immune response in rainbow trout [Oncorhynchus mykiss (Walbaum, 1792)] to experimental plerocercoid infections of Diphyllobothrium dendriticum (Nitzsch, 1824).

Development of the inflammatory response of rainbow trout to experimental infections with Diphyllobothrium dendriticum plerocercoids is described using light and electron microscopy. The cellular response to plerocercoids occurred within 2 weeks post-infection (p.i.). This was followed by an increase in leucocyte numbers during weeks 3-6 p.i., with full encapsulation of plerocercoids by week 6 p.i. Neutrophils were the first leucocytes to engage the developing plerocercoid, followed by large influxes of macrophages which transformed into epithelioid cells. With longer times p.i. the accumulation of different leucocyte types increased, and a blood vascular network developed. Full development of the composite cyst was characterized by fibroplasia, particularly at the periphery of the cyst, and the subsequent deposition of a collagenous tissue matrix. Enzyme-linked immunosorbent assay (ELISA) examination of serum samples taken over the 20 week period showed that specific anti-D. dendriticum antibody titres were first detected at 5 weeks p.i. and increased to a maximum by 11 weeks p.i.

[Differences in surface antigens of three diphyllobothriidae species, detected by serologic methods]. / Razlichiia poverkhnostnykh antigenov trekh vidov difillobotriid, vyiavliaemye serologicheskimi metodami.

Immunochemical studies of somatic and surface extracts from D. dendriticum, D. ditremum and D. latum tape-worms demonstrated low specificity of somatic antigens of all three species and serological specificity of D. latum. D. dendriticum and D. ditremum, though serologically close, contained specific components demanding more precise extraction methods.

Development of immunoreactivity to the invertebrate neuropeptide small cardiac peptide B in the tapeworm Diphyllobothrium dendriticum.

Neurons immunoreactive to small cardiac peptide B (SCPb) occur in the scolex and neck region of adult Diphyllobothrium dendriticum. The localisation of the SCPb-IR neurons in the peripheral nervous system is very pronounced; they are closely associated to the bothridial musculature in the scolex. SCPb-IR neurons were not observed in plerocercoid larvae but appeared after cultivation in vitro at 37 degrees C for 30 h. Functional and developmental aspects of the SCPb-IR neurons are discussed.

On the methods of obtaining diphyllobothriid surface antigens.

A new method of obtaining the diphyllobothriid surface antigens has been developed. It allows to obtain antigens with the increased content of the surface components of worm strobila. It was found that the antigens from extracts I are relatively more active than the antigens from extracts II and III.

[The agglutination test of coracidia of pseudophyllidean cestodes and its use in cestodological research]. / Reaktsiia aggliutinatsii koratsidiev psevdofillidnykh tsestod i ee ispol'zovanie v tsestodologicheskikh issledovaniiakh.

Ability of coracidia to agglutination in homologous antiserum was established. The method of reaction of a direct agglutination of coracidia (RDAC) in the culture of alive larvae was described. When comparing the RDAC results in homologous and heterologous variants a dependence of the agglutination pattern on the degree of antiserum homology was established. On the example of RDAC of the cestodes Diphyllobothrium dendriticum and D. ditremum in polyvalent and species-specific antisera to D. dendriticum possibility was shown of RDAC usage in the studies on specific taxonomy of pseudophyllid cestodes.

Coracidia agglutination reaction of pseudophyllid cestodes and its application in cestode studies.

The ability of cestode coracidia to agglutinate in homologous antiserum was established. The method of direct agglutination of coracidia (RDAC) in a culture of living larvae is described. The comparison of RDAC results in homologous and heterologous variants shows a dependence of the agglutination pattern on the degree of antiserum homology. By the example of RDAC of the cestodes Diphyllobothrium dentriticum and D. ditremum in polyvalent antisera and in antisera species-specific to D. dendriticum a possibility is shown of using RDAC in studies on species taxonomy of pseudophyllid cestodes.

[Antigenic affinity of species of the genus Diphyllobothrium (Cestoidea, Diphyllobothriidae) distributed in the territory of Karelia]. / Antigennoe srodstvo vidov roda Diphyllobothrium (Cestoidea, Diphyllobothriidae), rasprostranennykh na territorii Karelii.

Experiments on ringprecipitation reactions with homologous and heterologous antigenes of four species of diphyllobothriids have shown that D. dendriticum and D. ditremum are in a more close antigenic affinity. In relation to the above two species D. vogeli is approximately at the same level. D. latum displayed no antigenic affinity with the above species that, apparently, is connected with its parasitism only in mammals. Schemes of immunization of rabbits for obtaining antisera are given.

Activation of complement in C-reactive protein positive sera by phosphorylcholine-bearing component isolated from parasite extract.

Phosphorylcholine-bearing component levels in extracts of various parasites were determined by a capillary precipitin test using anti-phosphorylcholine Ig A myeloma protein. TEPC-15. Phosphorylcholine was demonstrated as a structural component not only in nematodes but also in trematodes and cestodes. The phosphorylcholine-bearing component was isolated from an extract of Toxocara canis larvae using a TEPC-15-Sepharose 4B column. The component reacted with C-reactive protein in sera to form one precipitin line in immunoelectrophoresis. The component provided two Brilliant Coomassie Blue positive bands in SDS-polyacrylamide gel electrophoresis. It reacted with C-reactive protein to activate complement in serum.