Studies on the host-parasite interaction between Diphyllobothrium spp. (Cestoda Pseudophyllidea) and rainbow trout, Oncorhynchus mykiss (Walbaum, 1792)
Diphyllobothrium dendriticum and D. ditremum have a wide distribution in the trout of Scottish lochs, although no specific trends in their overall distribution have been observed. The seasonal recruitment trends and development of infections with Diphyllobothrium spp. pleroceroids in wild rainbow trout, in one particular Scottish loch, were monitored regularly for the period 1986 to 1989. Infection varied between these years, but in 1987 intensities from March to November reached their highest levels in November when sampling ended. These two species reached infection intensities, in individual fish, higher than any previous reports and trout stocks appeared to be affected. Plerocercoids, in host-derived cysts, were located on the peritoneal stomach surface, the gross pathology of which is described. The cellular components of the host-derived cyst were examined in detail by light and electron microscopy. A number of leucocyte types, found within a collagenous matrix with associated fibroblasts, were observed. Leucocyte types included neutrophils, eosinophilic granular leucocytes, macrophages and occasional plasma cells. Antibody production, in response to natural infection, was examined by indirect immunofluorescence using serum on cryostat sections of plerocercoids obtained from wild-caught rainbow trout. The tegument of these larvae showed a positive fluorescence, indicating the presence of serum antibody in these trout. Semi-quantitative estimates of antibody titres were estimated by an optimised enzyme-linked immunosorbent assay (ELISA). To facilitate further examination of the events associated with the establishment of plerocercoids and associated host-derived cyst, under controlled conditions, a routinely maintained laboratory life cycle of D. dendriticum was established. This laboratory life cycle utilised Herring gulls (Larus agentatus) and the copepod, Cyclops abyssorum, as the definitive and first intermediate hosts respectively, and provided plerocercoid infections in trout which were examined at various times post infection. Tegumental extracts from the plerocercoids of D. dendriticum, obtained by freezing and thawing specimens, and conditioned medium obtained after in vitro maintenance of live plerocercoids, were prepared. These extracts were tested in respiratory burst assays and were found to stimulate the production of both hydrogen peroxide and superoxide anion by rainbow trout macrophages. The migratory responses of trout macrophages and neutrophils to these agents were also investigated, by an optimised Boyden technique using a 48 Well Micro Chemotaxis Chamber. Leucocytes were found to have an increased chemokinetic motility following stimulation/contact with these antigen preparations. Finally, to investigate if antigens on live parasites were attractive/stimulatory, an in vitro adherence assay was carried out. Procercoid stages, which share common antigens with the plerocercoid stage, were maintained in vitro and incubated with rainbow trout leucocyte suspensions, in the absence and presence of normal or immune serum from infected fish. Leucocytes adherence was considerably increased by the presence of immune serum, indicating the possible interaction of the non-specific and specific immune response in the host inflammatory reaction.